Mutagenesis of Active Site Residues in Type I Dehydroquinase from Escherichia coli

Name: Work Video:
Duration: 3 min 30 s
Andrew Leech; Richard James; John R. Coggins; Colin Kleanthous
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3097, 3146, 3175, 3180, 3226, 3290, 3298, 3354, 3374, 3385, 3483, 3560, 3639, 3642, 3667, 3674, 3715, 3723, 3751, 3754, 3817, 3831, 3842, 3869, 3961, 3973, 3987, 4034, 4073, 4149, 4156, 4161, 4163, 4183, 4263, 4298, 4338, 4379, 4393], 'type': [14, 314, 345, 645, 821, 824, 1006, 1109, 1343, 1443, 1790, 1930, 1963, 2044, 2050, 2128, 2342, 2673, 2676, 3123, 3376, 3884, 4006, 4083, 4209, 4265, 4273, 4300, 4389, 4448, 4455], 'I': [15, 315, 346, 646, 822, 1110, 1344, 1791, 1931, 2051, 2129, 2343, 2674, 2776], 'dehydroquinase': [16, 316, 347, 647, 771], 'from': [17, 257, 348, 588, 773, 1625, 1655, 2218, 2225, 2249, 2285, 2311, 2345, 2355, 2474, 2688, 3020, 3194, 3240, 3259, 3622, 3819, 3844, 4012], 'Escherichia': [18, 349, 774], 'coli.': [19, 350], 'To': [20, 351], 'further': [21, 352, 2038, 2238, 2571, 2908, 2961, 3369], 'test': [22, 353], 'their': [23, 354], 'importance,': [24, 355], 'these': [25, 356, 732, 1807, 2065, 2910, 3340], 'were': [27, 33, 358, 364, 884, 2070, 2229, 2328, 2373, 2402, 2433, 2502, 2567, 2681, 2718, 2755, 2770, 2882, 2890, 2900, 2912, 2998, 3086, 3104, 3116, 3128, 3189, 3192, 3203, 3238, 3250, 3258, 3410, 3454, 3465, 3493, 3499, 3508, 3513, 3534, 3562, 3647, 3756, 3761, 3888, 3954, 4010, 4038, 4068, 4187, 4201, 4244], 'mutated,': [28, 359], 'and': [29, 36, 67, 88, 172, 198, 210, 227, 360, 367, 398, 419, 503, 529, 541, 558, 690, 703, 726, 744, 823, 828, 1040, 1117, 1128, 1185, 1199, 1214, 1644, 1685, 1702, 1744, 1866, 1961, 2022, 2026, 2030, 2088, 2131, 2137, 2148, 2172, 2188, 2235, 2324, 2391, 2421, 2452, 2454, 2484, 2507, 2574, 2597, 2606, 2620, 2634, 2677, 2742, 2764, 2777, 2838, 2886, 2889, 2928, 3029, 3092, 3108, 3198, 3221, 3235, 3255, 3301, 3317, 3366, 3392, 3461, 3486, 3495, 3526, 3547, 3607, 3660, 3720, 3758, 3780, 3815, 3861, 3876, 3885, 3904, 3937, 3952, 4007, 4022, 4054, 4063, 4171, 4240, 4249, 4257, 4266, 4415, 4461], 'resulting': [31, 50, 362, 381, 1279, 2033, 2462, 3114, 3463, 3497], 'mutants': [32, 363, 3950, 4186, 4251], 'expressed,': [34, 365], 'purified,': [35, 366], 'characterized.': [37, 368], 'When': [38, 369], 'highly': [40, 371], 'conserved,': [41, 116, 372, 447], 'Schiff': [42, 201, 225, 276, 373, 532, 556, 607, 1209, 1529, 2159, 2986], 'base-forming': [43, 374, 2160], 'lysine': [44, 375, 2161], 'residue': [45, 119, 181, 376, 450, 512, 1637, 1696, 2087, 2169], 'was': [46, 60, 377, 391, 1697, 1710, 1714, 1862, 2083, 2109, 2155, 2181, 2201, 2217, 2224, 2272, 2284, 2309, 2348, 2444, 2464, 2479, 2518, 2522, 2539, 2558, 2589, 2800, 2810, 2848, 2867, 2954, 3063, 3140, 3232, 3273, 3294, 3342, 3382, 3431, 3575, 3580, 3594, 3638, 3679, 3729, 3783, 3899, 3918, 3928, 3939, 4002, 4167], 'mutated': [47, 378, 2018, 2110, 2136], '(K170A)': [48, 379], 'enzyme': [51, 266, 326, 382, 597, 657, 772, 1008, 1178, 1684, 1814, 1840, 1988, 2144, 2887, 2971, 3124, 3196, 3356, 3377, 4390, 4449, 4456], 'showed': [52, 383, 4211], 'a': [53, 72, 83, 93, 158, 261, 275, 384, 403, 414, 424, 489, 592, 606, 869, 890, 932, 1051, 1208, 1449, 1578, 1583, 1609, 1634, 1890, 1929, 1962, 2211, 2273, 2331, 2376, 2392, 2405, 2411, 2511, 2570, 2685, 2852, 2870, 2932, 3018, 3027, 3206, 3212, 3223, 3281, 3468, 3502, 3516, 3531, 3650, 3661, 3896, 3921, 4181, 4212, 4216, 4295], '˜106-fold': [54, 385], 'reduction': [55, 386, 1223, 1267, 3666], 'catalytic': [57, 153, 388, 484, 3352], 'activity,': [58, 389], 'but': [59, 294, 390, 625, 877, 922, 1816, 1836, 1913, 2662, 3977], 'still': [61, 392], 'able': [62, 287, 393, 618, 3955], 'to': [63, 175, 270, 288, 292, 394, 506, 601, 619, 623, 687, 809, 880, 1540, 1639, 1651, 1820, 2024, 2028, 2036, 2076, 2090, 2098, 2139, 2162, 2363, 2379, 2551, 2649, 2802, 2851, 2902, 2921, 2931, 3121, 3186, 3211, 3242, 3261, 3334, 3397, 3412, 3434, 3447, 3452, 3467, 3490, 3501, 3544, 3555, 3582, 3767, 3871, 3956, 3995, 4076, 4147, 4173, 4195, 4418], 'bind': [64, 170, 395, 501], 'both': [65, 396, 1795, 1928, 2624, 3882], 'substrate': [66, 108, 291, 397, 439, 622, 1213, 3283, 3350], 'product,': [68, 272, 399, 603, 3302], 'as': [69, 137, 182, 400, 468, 513, 738, 796, 804, 887, 1015, 1725, 1884, 2057, 3026, 3110, 3153, 3523, 3530, 3682, 3731, 3941, 3997, 4018, 4279, 4285, 4291, 4317, 4344, 4405, 4450], 'shown': [70, 401, 1193, 1265, 1435, 2138, 4172], 'by': [71, 239, 324, 402, 570, 655, 1113, 1222, 1441, 1736, 1832, 1864, 2008, 2183, 2203, 2358, 2435, 2447, 2470, 2576, 2684, 2692, 2758, 2772, 2812, 2884, 2958, 3001, 3030, 3088, 3094, 3125, 3130, 3142, 3148, 3315, 3344, 3456, 3684, 3826, 3859, 3902, 4154, 4169, 4190, 4356], 'novel': [73, 404], 'fluorescence-based': [74, 405], 'ligand-binding': [75, 406], 'assay.': [76, 407], 'This': [77, 408, 1861, 2520], 'is': [78, 100, 192, 248, 255, 286, 302, 409, 431, 523, 579, 586, 617, 633, 691, 766, 1042, 1192, 1337, 1423, 1448, 1493, 1785, 1797, 1841, 2113, 2124, 2428, 3280, 3847, 3931, 4137], 'consistent': [79, 410, 1424, 1604], 'with': [80, 96, 164, 260, 328, 411, 427, 495, 591, 659, 868, 1211, 1224, 1425, 1582, 1605, 1834, 2207, 2375, 2450, 2504, 2560, 2596, 2612, 2627, 2640, 2744, 2774, 2805, 2869, 3022, 3351, 3549, 3601, 3763, 4215, 4221, 4304, 4312, 4413, 4459], 'Lys-170': [81, 412, 1206, 1336, 2023], 'playing': [82, 413], 'central': [84, 415, 697, 1427], 'role': [85, 178, 307, 416, 509, 638, 1428], 'catalysis': [87, 418, 2171], 'shows': [89, 148, 157, 420, 479, 488], 'that,': [90, 421, 1436], 'although': [91, 422], 'forming': [92, 423], 'covalent': [94, 425, 1058], 'bond': [95, 426], 'substrate,': [98, 429, 3299], 'it': [99, 191, 430, 522, 1041, 1520, 1709, 2112, 3430], 'not': [101, 432, 1130, 1810, 3395, 4178], 'essential': [102, 433, 2096], 'for': [103, 114, 146, 434, 445, 477, 741, 893, 1004, 1189, 1498, 1570, 1799, 1813, 1908, 2072, 2157, 2481, 2569, 2582, 2748, 2782, 2793, 3065, 3287, 3307, 3407, 3418, 3585, 3627, 3634, 3773, 3794, 3808, 3933, 3945, 4061, 4080, 4246, 4383, 4397], 'ground': [104, 435, 2173], 'state': [105, 436, 2174], 'binding': [106, 437, 2175, 3292, 3471, 3775], 'or': [109, 126, 135, 440, 457, 466, 803, 1700, 2233, 3538, 3603, 3802, 4044, 4308], 'product.': [110, 441], 'Conversely,': [111, 442], 'substituting': [112, 443], 'leucine': [113, 444, 2029], 'iodoacetate-reactive': [117, 448], 'methionine': [118, 449, 1741, 2121], '(M205L)': [120, 451], 'had': [121, 130, 452, 461, 3359, 4026], 'little': [122, 453, 4028], 'effect': [123, 454, 2142, 4029], 'on': [124, 455, 1274, 1703, 2143, 2330, 2516, 2541, 2623, 2659, 3205, 3477, 3515, 3649, 3967, 3999, 4015, 4030, 4052, 4055], 'kcat': [125, 456], 'Km.': [127, 458], 'Diethylpyrocarbonate': [128, 459], 'either': [133, 464, 797, 1698, 1909, 4305], 'His-143': [134, 309, 465, 640, 1699, 1713], 'His-146': [136, 467], 'putative': [139, 470, 1716], 'active': [140, 471, 811, 1822, 2614, 2629, 3355, 3672, 3949], 'site': [141, 472, 2384, 2394, 2413, 2417, 3069, 3673], 'general': [142, 185, 473, 516, 1610, 1717], 'base.': [143, 474], 'Substituting': [144, 475], 'alanine': [145, 476, 2025, 2099], 'each': [147, 478, 3628, 3635, 4162], 'that': [149, 190, 214, 228, 251, 282, 295, 480, 521, 545, 559, 582, 613, 626, 1044, 1205, 1266, 1524, 1692, 1712, 1806, 2654, 2689, 3577, 3853, 3898, 4140], 'H146A': [150, 481, 4403, 4409], 'retains': [151, 482], 'full': [152, 483], 'activity': [154, 485, 1735, 1815, 2888, 3139], 'while': [155, 486], 'H143A': [156, 168, 216, 253, 284, 487, 499, 547, 584, 615, 2645, 2965, 3404, 3479, 3989, 4277, 4339], '106-fold': [159, 490], 'loss': [160, 491, 2437], 'activity.': [162, 493, 2145, 2440], 'As': [163, 494, 4180], 'K170A': [166, 497, 3980, 4248, 4283], 'mutant,': [167, 498, 3405, 3480, 3990], 'can': [169, 500, 1219, 1829], 'ligand,': [171, 502], 'addition': [174, 505, 4378, 4392], 'predicted': [177, 508], 'this': [180, 229, 246, 325, 511, 560, 577, 656, 1269, 1620, 1648, 2014, 2168, 2537, 2897, 2952, 3428, 3845, 3879], 'proton-abstracting': [184, 515], 'base,': [186, 226, 517, 557], 'our': [187, 518], 'data': [188, 280, 519, 611, 1916, 1997, 3464, 3498], 'suggest': [189, 281, 520, 612], 'also': [193, 524, 1263, 3432, 4188], 'involved': [194, 525, 730, 799, 2967], 'formation': [197, 221, 528, 552, 1526], 'breakdown': [199, 530], 'base': [202, 277, 533, 608, 1210, 1530, 1611, 1621, 1718], 'intermediates.': [203, 534], 'Isoelectric': [204, 535], 'focusing,': [205, 536], 'electrospray': [206, 537, 3591], 'ionization': [207, 538, 676], 'mass': [208, 539, 677, 874, 1127, 3132, 3625, 3636], 'spectrometry,': [209, 540], 'fluorescence': [211, 542, 3400], 'spectroscopy': [212, 543], 'show': [213, 544], 'mutant': [217, 254, 285, 548, 585, 616, 2034, 2529, 2646, 2678, 3102, 3429, 3886, 3894, 3927, 3981, 4008, 4165, 4267], 'preferentially': [218, 549], 'stabilizes': [219, 550], 'product': [224, 293, 301, 555, 624, 632, 2988, 3147, 3668, 3724], 'results': [230, 561], 'burst': [232, 563], 'kinetics': [233, 564], 'reminiscent': [234, 565], 'p-nitrophenyl': [236, 567], 'acetate': [237, 568], 'hydrolysis': [238, 569], 'chymotrypsin.': [240, 571], 'The': [241, 304, 572, 635, 706, 712, 760, 1000, 1108, 1175, 1617, 2053, 2067, 2280, 2371, 2398, 2441, 2461, 2565, 2644, 2753, 2762, 2808, 2845, 3101, 3113, 3229, 3337, 3371, 3630, 3840, 4024], 'most': [242, 573], 'striking': [243, 574], 'illustration': [244, 575], 'stabilization': [247, 578], 'fact': [250, 581], 'isolated': [256, 587, 1689, 2446], 'overexpressing': [258, 589], 'cells': [259, 590, 2566, 2754], 'significant': [262, 593], 'proportion': [263, 594], 'monomers': [267, 598], 'covalently': [268, 599, 3725, 4313], 'bound': [269, 600, 3726, 4314], '3-dehydroshikimate,': [273, 604], 'via': [274, 605], 'linkage.': [278, 609], 'Our': [279, 610], 'slowly': [289, 620, 2664], 'transform': [290, 621], 'hydrolytic': [297, 628], 'release': [298, 629], 'stalled.': [303, 634], 'proposed': [305, 636], 'dual': [306, 637], 'may': [317, 648, 1809, 1817], 'explain': [318, 649], 'why': [319, 650], 'elimination': [321, 652, 1046, 1492], 'reaction': [322, 653, 1439, 1447], 'catalyzed': [323, 654, 1440], 'proceeds': [327, 658], 'syn': [329, 660, 1450], 'stereochemistry.': [330, 661], 'INTRODUCTIONDehydroquinase': [662], '(DHQase;': [663], '3-dehydroquinate': [664, 2180, 2206], 'dehydratase)': [665], '1The': [666], 'abbreviations': [667], 'used': [668, 2236, 3064, 3367, 3563, 3940, 4060], 'are:': [669], 'DHQase3-dehydroquinaseDHQ3-dehydroquinic': [670], 'acidDHS3-dehydroshikimic': [671], 'acidPCRpolymerase': [672], 'chain': [673], 'reactionIEFisoelectric': [674], 'focusingESI-MSelectrospray': [675], 'spectrometryPAGEpolyacrylamide': [678], 'gel': [679, 2935, 4193], 'electrophoresis.': [680], 'catalyses': [681], 'dehydration': [683, 1003], '3-dehydroquinic': [685], '3-dehydroshikimic': [688], 'third': [693], 'step': [694], 'shikimate': [698, 929], 'pathway': [699, 717, 892, 930], 'microorganisms,': [701], 'fungi,': [702], 'plants(1Haslam': [704], 'E.': [705, 1115, 1176, 1571, 1731, 2289, 2317, 2346, 2424, 2467, 2531, 3308, 3923, 3962], 'Shikimate': [707], 'Pathway.': [708], 'Butterworths,': [709], 'London1974Google': [710], 'Scholar).': [711, 759, 861, 999, 1107, 1174, 1260, 1335, 1489, 1678, 1778, 1860, 2198, 2712, 3084, 3172, 3710, 3749, 3916, 4135, 4335], 'end': [713, 2015], 'products': [714], 'are': [718, 813, 865, 1121, 1129, 1904, 4145], 'precursors': [720], 'aromatic': [723, 2599, 2641, 2666, 3934, 3974], 'acids,': [725], 'so': [727, 3393, 3938], 'enzymes': [729, 867, 2680], 'biosynthetic': [733], 'reactions': [734], 'been': [736, 788, 831, 925, 1013, 1182, 1264, 1522, 1623, 1990, 2135, 3313, 3360], 'studied': [737, 1183], 'potential': [739], 'sites': [740], 'antimicrobial': [742], 'agents': [743], 'herbicides(2Kishore': [745], 'G.M.': [746], 'Shah': [747], 'D.M.': [748], 'Annu.': [749], 'Rev.': [750], 'Biochem.': [751, 852, 945, 1078, 1139, 1165, 1251, 1362, 1380, 1394, 1594, 2193, 2259, 2703, 3008, 3163, 4102], '1988;': [752], '57:': [753], '627-663Crossref': [754], 'PubMed': [755, 857, 917, 950, 974, 995, 1083, 1144, 1170, 1256, 1301, 1333, 1367, 1385, 1399, 1417, 1485, 1515, 1599, 1676, 1760, 1776, 1858, 1881, 1956, 1983, 2264, 2306, 2708, 3012, 3057, 3080, 3168, 3329, 3708, 3914, 4107, 4133, 4442, 4483], 'Scopus': [756, 858, 918, 951, 975, 996, 1036, 1084, 1145, 1171, 1239, 1257, 1368, 1386, 1400, 1418, 1486, 1516, 1600, 1957, 1984, 2265, 2709, 3013, 3081, 3169, 3330, 3746, 4108, 4236, 4332], '(350)': [757], 'Google': [758, 860, 920, 953, 977, 998, 1038, 1086, 1106, 1147, 1173, 1241, 1259, 1302, 1334, 1370, 1388, 1402, 1420, 1469, 1488, 1518, 1564, 1602, 1677, 1761, 1777, 1859, 1882, 1959, 1986, 2267, 2307, 2711, 3015, 3058, 3083, 3171, 3332, 3709, 3748, 3915, 4110, 4134, 4238, 4334, 4443, 4484], 'focus': [761], 'present': [764], 'work': [765], 'coli': [775, 1116, 1177, 1572, 1732, 2290, 2318, 2347, 2425, 2468, 2532, 3309, 3924, 3963], '(EC': [776], '4.2.1.10).': [777], 'In': [778, 1992], 'particular,': [779], 'roles': [781, 1999, 2040, 2063], 'several': [783, 2056], 'investigated': [789], 'which': [790, 878, 1422, 1630, 2079, 2123, 2409, 2984], 'chemical': [791, 2009], 'being': [798, 805], 'close': [807, 1819, 1887], 'proximity': [808, 1888], 'site.There': [812], 'two': [814, 1738, 1781, 2073, 2082, 2119, 3721, 4309], 'distinct': [815], 'classes': [816], 'DHQase': [818, 1345, 1573, 2208, 2344, 2679, 3279, 3310, 3675, 3716, 4210, 4278, 4284, 4290, 4301, 4404, 4410], 'enzymes,': [819, 1120], 'designated': [820], 'II,': [825], 'whose': [826], 'properties': [827, 2093, 4033], 'mechanisms': [829], 'compared': [832], '(3Kleanthous': [834, 1060], 'C.': [835, 1021, 1061, 1091, 1097, 1281, 1323, 1454, 1460, 1549, 1555, 1664, 1746, 1764, 1846, 1869, 1947, 3045, 3688, 3738, 4085, 4113, 4228, 4324, 4422, 4463], 'Deka': [836, 1062, 1155, 1352, 1944, 4086], 'R.': [837, 1063, 1156, 1353, 1393, 1405, 1662, 1945, 2150, 2190, 3043, 3074, 4087], 'Davis': [838, 1064, 1308, 3318, 3321, 4088], 'K.': [839, 1065, 1246, 1309, 1375, 2252, 2278, 4089], 'Kelly': [840, 1066, 1161, 1286, 1310, 1358, 3693, 4090, 4118, 4427, 4468], 'S.M.': [841, 1067, 1162, 1311, 1359, 1935, 4091], 'Cooper': [842, 1068, 1159, 1284, 1356, 3691, 4092, 4116, 4425, 4466], 'A.': [843, 913, 1017, 1069, 1160, 1285, 1305, 1357, 1481, 1511, 3692, 3734, 4093, 4117, 4224, 4320, 4426, 4467], 'Harding': [844, 1070, 1316, 4094], 'S.E.': [845, 1071, 1317, 4095], 'Price': [846, 1072, 1163, 1288, 1318, 1360, 3695, 4096, 4120, 4429, 4470], 'N.C.': [847, 1073, 1164, 1289, 1319, 1361, 3696, 4097, 4121, 4430, 4471], 'Hawkins': [848, 959, 1024, 1074, 1094, 1151, 1348, 1457, 1552, 1942, 1975, 4098], 'A.R.': [849, 960, 1025, 1075, 1095, 1152, 1349, 1458, 1553, 1943, 1976, 4099], 'Coggins': [850, 943, 985, 1022, 1076, 1092, 1137, 1157, 1249, 1290, 1320, 1354, 1378, 1410, 1455, 1550, 1592, 1665, 1749, 1765, 1847, 1867, 1870, 1948, 1967, 2257, 2701, 3046, 3161, 3697, 3735, 4100, 4122, 4225, 4321, 4431, 4472], 'J.R.': [851, 944, 986, 1023, 1077, 1093, 1138, 1158, 1250, 1291, 1321, 1355, 1379, 1411, 1456, 1551, 1593, 1666, 1750, 1766, 1848, 1871, 1949, 1968, 2258, 2702, 3047, 3162, 3698, 3736, 4101, 4123, 4226, 4322, 4432, 4473], 'J.': [853, 938, 946, 989, 1019, 1079, 1089, 1098, 1140, 1166, 1232, 1252, 1292, 1313, 1324, 1363, 1381, 1395, 1452, 1461, 1547, 1556, 1595, 1667, 1751, 1767, 1849, 1872, 1950, 1977, 2149, 2260, 2298, 2301, 2704, 3048, 3164, 3699, 3906, 3909, 4103, 4124, 4433, 4474], '1992;': [854, 992, 1080, 1670, 1953, 3051, 4104], '282:': [855, 1081, 4105], '687-695Crossref': [856, 1082, 4106], '(84)': [859, 1085, 4109], 'Type': [862], 'II': [863, 1007, 1444, 1964], 'DHQases': [864, 3887], 'thermostable': [866], 'relatively': [870, 4027], 'small': [871], 'subunit': [872], 'molecular': [873, 1126], '(˜16': [875], 'kDa)': [876], 'oligomerize': [879], 'form': [881], 'dodecamers.': [882], 'They': [883, 4048], 'originally': [885], 'identified': [886, 1014, 2007], 'part': [888], 'catabolic': [891], 'utilization': [894], 'quinic': [896], 'fungi(4Giles': [899], 'N.H.': [900], 'Partridge': [901], 'C.W.H.': [902], 'Ahmed': [903], 'S.I.': [904], 'Case': [905], 'M.E.': [906], 'Proc.': [907, 1475, 1505], 'Natl.': [908, 1476, 1506], 'Acad.': [909, 1477, 1507], 'Sci.': [910, 1478, 1508], 'U.': [911, 1479, 1509], 'S.': [912, 958, 1132, 1244, 1287, 1373, 1480, 1510, 1587, 2254, 2696, 3156, 3320, 3694, 4119, 4428, 4469], '1967;': [914], '58:': [915], '1930-1937Crossref': [916], '(36)': [919], 'Scholar),': [921, 1039, 1421, 1519, 1603, 3016, 4239], 'since': [924, 1924, 2111, 3920], 'found': [926, 3852], 'number': [933], 'prokaryotes(5White': [935], 'P.J.': [936], 'Young': [937, 963], 'Hunter': [939], 'I.S.': [940], 'Nimmo': [941], 'H.G.': [942], '1990;': [947, 1754, 1770, 1852, 1875], '265:': [948, 1755, 1771, 1853, 1876], '735-738Crossref': [949], '(50)': [952], 'Scholar,': [954, 978, 1087, 1148, 1242, 1303, 1371, 1389, 1403, 1470, 1762, 4111], '6Garbe': [955], 'T.': [956], 'Servos': [957], 'Dimitriadis': [961], 'G.': [962], 'D.': [964], 'Charles': [965, 1153, 1350, 1940, 1973], 'I.G.': [966, 1154, 1351, 1941, 1974], 'Mol.': [967, 1951, 1978, 3075], '&': [968, 1029], 'Gen.': [969, 990], 'Genet.': [970], '1991;': [971, 1253, 1295, 1382, 3077, 3702, 4127, 4436, 4477], '228:': [972], '385-392Crossref': [973], '(75)': [976], '7Euverink': [979], 'G.J.W.': [980], 'Hessels': [981], 'G.J.': [982], 'Vrijbloed': [983], 'J.W.': [984], 'Dijkhuizen': [987], 'L.': [988, 1937], 'Microbiol.': [991, 3076], '138:': [993], '2449-2457Crossref': [994], '(34)': [997, 1985], 'stereochemistry': [1001], 'Anacystis': [1010], 'nidulans': [1011], 'has': [1012, 1181, 1262, 1271, 1521, 1622, 2133, 2410, 3311], 'anti(8Shneier': [1016], 'Harris': [1018], 'Kleanthous': [1020, 1090, 1322, 1453, 1548, 1663, 1865, 1946, 3044, 3685, 3737, 4227, 4323, 4419], 'Abell': [1026, 1096, 1459, 1554], 'C.A.': [1027, 1391], 'Bioorg.': [1028, 3739, 4229, 4325], 'Med.': [1030, 3740, 4230, 4326], 'Chem.': [1031, 1099, 1101, 1234, 1294, 1326, 1462, 1464, 1557, 1559, 1669, 1753, 1769, 1851, 1874, 3050, 3701, 3741, 4126, 4231, 4327, 4435, 4476], 'Lett.': [1032, 1413, 3742, 4232, 4328], '1993;': [1033, 1103, 1167, 1364, 1466, 1561, 3743, 4233, 4329], '3:': [1034, 3744, 4234, 4330], '1399-1402Crossref': [1035], '(35)': [1037], 'thought': [1043], 'water': [1048, 3537], 'occurs': [1049], 'through': [1050], 'concerted': [1052], 'without': [1054, 2237, 2598, 2665, 3368, 4041], 'involvement': [1056], 'intermediates': [1059], '9Harris': [1088], 'Soc.': [1100, 1235, 1463, 1558], 'Commun.': [1102, 1465, 1560], '1993:': [1104, 1467, 1562], '1080-1081Crossref': [1105, 1468, 1563], 'class,': [1111, 1445], 'typified': [1112], 'Salmonella': [1118], 'typhi': [1119], 'dimers': [1122, 4303], '25-kDa': [1124], 'monomer': [1125], 'heat-stable(10Chaudhuri': [1131], 'Lambert': [1133, 1588, 2697, 3157], 'J.M.': [1134, 1589, 2698, 3158], 'McColl': [1135, 1590, 2699, 3159], 'L.A.': [1136, 1591, 2700, 3160], '1986;': [1141, 1596, 2261, 2705, 3165], '239:': [1142, 1597, 2706, 3166], '699-704Crossref': [1143, 1598, 2707, 3167], '(47)': [1146, 1601, 2710, 3170], '11Moore': [1149], 'J.D.': [1150, 1347, 1939, 1972], '295:': [1168, 1365], '277-285Crossref': [1169, 1366], '(32)': [1172, 1369, 1401], 'particular': [1180], 'extensively,': [1184], 'current': [1187], 'model': [1188], 'its': [1190, 1426], 'Fig.': [1195, 1722], 'S1.': [1196], 'peptide': [1200, 1690], 'mapping': [1201, 1680], 'studies': [1202, 3341], 'established': [1204, 1691], 'forms': [1207], 'product;': [1215], 'latter': [1217], 'intermediate': [1218, 1270, 2651], 'be': [1220, 1811, 1830, 3335, 4174], 'trapped': [1221], 'sodium': [1225, 3677], 'borohydride(12Butler': [1226], 'R.J.': [1227, 3788], 'Alworth': [1228], 'W.L.': [1229], 'Nugent': [1230], 'M.J.': [1231], 'Am.': [1233], '1974;': [1236], '96:': [1237], '1617-1618Crossref': [1238], '(66)': [1240], '13Chaudhuri': [1243, 1372], 'Duncan': [1245, 1374], 'Graham': [1247, 1376], 'L.D.': [1248, 1377], '275:': [1254, 1383], '1-6Crossref': [1255, 1384], '(58)': [1258, 1387], 'It': [1261, 4136], 'profound': [1272], 'effects': [1273, 4043, 4143], 'stability': [1276], 'protein(14Kleanthous': [1280], 'Reilly': [1282, 3689, 4114, 4423, 4464], 'M.': [1283, 3006, 3690, 4115, 4424, 4465], 'Biol.': [1293, 1325, 1668, 1752, 1768, 1850, 1873, 1952, 1979, 3049, 3700, 4125, 4434, 4475], '266:': [1296, 3703, 4128, 4437, 4478], '10893-10898Abstract': [1297, 3704, 4129, 4438, 4479], 'Full': [1298, 1330, 1673, 1757, 1773, 1855, 1878, 3054, 3705, 4130, 4439, 4480], 'Text': [1299, 1331, 1674, 1758, 1774, 1856, 1879, 3055, 3706, 4131, 4440, 4481], 'PDF': [1300, 1332, 1675, 1759, 1775, 1857, 1880, 3056, 3707, 4132, 4441, 4482], '15Reilly': [1304], 'Morgan': [1306], 'P.': [1307], 'Greene': [1312], 'Rowe': [1314], 'A.J.': [1315], '1994;': [1327, 1396, 1414, 1980], '269:': [1328], '5523-5526Abstract': [1329], 'conserved': [1338, 1787, 2125], 'all': [1340, 2001, 3117, 4037, 4049], 'known': [1342, 2105], 'sequences(11Moore': [1346], '16Bonner': [1390], 'Jensen': [1392], '302:': [1397], '11-14Crossref': [1398], '17Deka': [1404], 'Anton': [1406], 'I.A.': [1407, 1474, 1504], 'Dunbar': [1408], 'B.': [1409, 2313], 'FEBS': [1412], '349:': [1415], '397-402Crossref': [1416], '(26)': [1419], 'mechanism.': [1431, 2106], 'Stereochemical': [1432], 'unlike': [1437], 'elimination(9Harris': [1451], '18Hanson': [1471], 'K.R.': [1472, 1502], 'Rose': [1473, 1503], '1963;': [1482, 1512], '50:': [1483, 1513], '981-988Crossref': [1484, 1514], '(86)': [1487, 1517], 'Since': [1490, 3278], 'anti': [1491], 'chemically': [1494], 'preferred': [1496], 'course': [1497, 3475, 3569, 3778], 'uncatalyzed': [1500], 'reaction(18Hanson': [1501], 'suggested': [1523], 'involves': [1531], 'some': [1532], 'distortion': [1533], 'carbocyclic': [1536], 'ring': [1537], 'dehydroquinate': [1539, 3181, 4380, 4394, 4414, 4460], 'render': [1541], 'pro-R': [1543], 'proton': [1544], 'more': [1545, 2892, 3604], 'reactive(9Harris': [1546], 'Scholar).The': [1565], 'pH': [1566, 2735, 2836, 2918, 2978, 3268, 3542, 4372], 'dependence': [1567], 'Vmax': [1569], 'reveals': [1574], 'presence': [1576, 2546], 'single': [1579, 1635, 3089, 3282, 3469, 3503, 4213], 'ionizing': [1580], 'species': [1581], 'pKa': [1584, 1646, 1653], '6.2(10Chaudhuri': [1586], 'action': [1607], '(Fig.': [1615, 4252], 'S1).': [1616], 'identity': [1618], 'inferred': [1624], 'diethyl': [1626], 'pyrocarbonate': [1627], 'histidine': [1636, 1695, 2068, 2097], 'led': [1638], 'inactivation': [1640, 1801, 4412, 4458], 'enzyme,': [1643, 1804, 3284], 'corresponded': [1650], 'determined': [1654, 3314, 4245], 'pH-Vmax': [1657], 'profiles': [1658], 'enzyme(19Deka': [1661], '267:': [1671, 3052], '22237-22242Abstract': [1672, 3053], 'Peptide': [1679], 'modified': [1683, 2687, 3060], 'sequencing': [1686, 2325, 2489, 3091, 3096], 'an': [1688, 1726, 2381, 2415, 2477, 2495, 2552, 2650, 2925, 3295, 3942], 'reactive': [1694, 1740, 2120], 'His-146,': [1701, 2021], 'basis': [1705], 'sequence': [1707, 2377], 'comparisons': [1708], 'concluded': [1711], 'B': [1719], 'highlighted': [1720], 'S1.Iodoacetate': [1723], 'acts': [1724], 'affinity': [1727], 'labeling': [1728], 'reagent': [1729, 2231], 'DHQase,': [1733, 1912], 'destroying': [1734], 'alkylating': [1737], 'unusually': [1739], 'residues,': [1742, 1782], 'Met-23': [1743, 2132], 'Met-205(20Kleanthous': [1745], 'Campbell': [1747, 2255], 'D.G.': [1748, 1966], '10929-10934Abstract': [1756], '21Kleanthous': [1763], '10935-10939Abstract': [1772, 1854, 1877], 'Of': [1779], 'only': [1783, 1837, 2115, 2637, 3991], 'Met-205': [1784, 1828, 2027, 2107], 'completely': [1786], 'family.': [1792], 'Alkylation': [1793], 'required': [1798], 'complete': [1800, 3095], 'implying': [1805], 'crucial': [1812], 'lie': [1818], 'site.': [1823], 'Moreover,': [1824], 'carboxymethylation': [1826], 'reversed': [1831], 'treatment': [1833, 2204, 2962], '2-mercaptoethanol': [1835], 'when': [1838, 2638, 3965], 'folded': [1844], 'state(21Kleanthous': [1845], 'interpreted': [1863], '(21Kleanthous': [1868], 'Scholar)': [1883, 1960, 1987, 2268, 2308, 3333, 3801, 3813], 'demonstrating': [1885], 'neutralizing': [1891], 'positive': [1892], 'charge': [1893, 4199], 'near': [1894, 1922], 'carboxylate': [1896], 'moiety': [1897], 'carboxymethylated': [1900], 'Met-205.At': [1901], 'present,': [1902], 'there': [1903], 'no': [1905, 2141, 2628], 'three-dimensional': [1906], 'structures': [1907], 'class': [1910], 'high': [1914], 'resolution': [1915], 'should': [1917], 'become': [1918], 'available': [1919], 'future,': [1923], 'diffracting': [1925], 'crystals': [1926], '(22Boys': [1932], 'C.W.G.': [1933], 'Bury': [1934], 'Sawyer': [1936], 'Moore': [1938, 1971], '227:': [1954], '352-355Crossref': [1955], '(14)': [1958], '(23Gourley': [1965], 'Isaacs': [1969], 'N.W.': [1970], '241:': [1981], '488-491Crossref': [1982], 'produced.': [1991], 'absence': [1994, 3972], 'such': [1996, 3576], 'acids': [2004, 2601, 3936, 3976], 'thus': [2005], 'far': [2006], 'need': [2011], 'clarification.': [2012], 'Toward': [2013], 'we': [2016, 3851], 'His-143,': [2020], 'characterized': [2031], 'proteins': [2035, 3115, 4009, 4166], 'elucidate': [2037], 'wild': [2043, 2675, 3122, 3375, 3883, 4082, 4264, 4272, 4299, 4388, 4447, 4454], 'DHQase.': [2052, 3947, 4340], 'mutations': [2054, 2069, 4025], 'addressed': [2055], 'yet': [2058], 'unanswered': [2059], 'questions': [2060], 'concerning': [2061], 'residues.': [2066], 'made': [2071], 'reasons,': [2074], 'first': [2075, 2690, 3900], 'determine': [2077, 4196], 'unambiguously': [2078], 'likely': [2085], 'DEPC-reactive': [2086], 'second': [2089], 'assess': [2091, 2163], 'mutation': [2100], 'terms': [2102], 'alone': [2108], 'one': [2116, 2508], 'family': [2130], 'already': [2134], '2E.': [2146], 'Borthwell': [2147], 'Coggins,': [2151], 'unpublished': [2152], 'observations.': [2153], 'Alanine': [2154], 'substituted': [2156], 'relative': [2165], 'importance': [2166], 'ligand.MATERIALS': [2177], 'AND': [2178], 'METHODSChemicalsAmmonium': [2179], 'prepared': [2182, 2202, 2951, 3730, 4316], 'method': [2185, 3003, 3062], 'Grewe': [2187], 'Haendler(24Grewe': [2189], 'Haendler': [2191], 'H.': [2192], 'Prep.': [2194], '1968;': [2195], '12:': [2196], '21-26Google': [2197], 'Ammonium': [2199], '3-dehydroshikimate': [2200], '(this': [2209], 'produces': [2210], '15:1': [2212], 'dehydroshikimate:dehydroquinate': [2213], 'mixture).': [2214], 'Hydroxylamine': [2215], 'hydrochloride': [2216, 4075, 4354], 'Sigma.': [2219], 'DE52': [2220, 2856], 'ion': [2221, 4057], 'exchange': [2222, 2858, 4058], 'resin': [2223, 2859], 'Whatman.': [2226], 'Other': [2227], 'chemicals': [2228], 'grade': [2232], 'better': [2234], 'purification.Genetic': [2239], 'MaterialsPlasmid': [2240], 'pKD203': [2241, 2357], '(which': [2242, 2427], 'contains': [2243, 3821], '1.8-kilobase': [2245], 'pair': [2246, 3830], 'genomic': [2247], 'fragment': [2248, 2443], 'pKD201': [2250], '(25Duncan': [2251], 'Chaudhuri': [2253, 2693], 'M.S.': [2256], '238:': [2262], '475-483Crossref': [2263], '(40)': [2266], 'ligated': [2269, 2403, 2455, 3105], 'into': [2270, 2404, 2423, 2456, 2466, 2525, 3106, 3864, 3890], 'pKK223-3)': [2271], 'kind': [2274], 'gift': [2275], 'Dr.': [2277, 2312], 'Duncan.': [2279], 'expression': [2281, 2458, 2557, 3837, 3843, 3870, 3880], 'vector': [2282, 2408, 2459, 2633, 3838], 'pKK223-3': [2283, 3107], 'Pharmacia': [2286, 3651, 3662], 'Biotech': [2287], 'Inc.': [2288], 'strain': [2291, 3897], 'AB2848': [2292, 2588, 3917, 3930, 3964, 4013], '(tsx-356,': [2293], 'supE42,': [2294], 'λ-,': [2295], 'aroD352)': [2296], '(26Pittard': [2297], 'Wallace': [2299, 3907], 'B.J.': [2300, 3908], 'Bacteriol.': [2302, 3910], '1966;': [2303, 3911], '91:': [2304, 3912], '1494-1508Crossref': [2305, 3913], 'obtained': [2310], 'Bachmann': [2314], 'at': [2315, 2385, 2395, 2578, 2751, 2785, 2789, 2796, 2877, 2896, 3039, 3150, 3199, 3270, 3275, 3347, 3363, 3415, 3419, 3423, 3670, 4070], 'Genetic': [2319], 'Stock': [2320], 'Center.': [2321], 'Mutagenic,': [2322], 'PCR,': [2323], 'primer': [2326], 'oligonucleotides': [2327], 'synthesized': [2329], 'New': [2332], 'Brunswick': [2333], 'Cyclone': [2334], 'synthesizer.Overexpression': [2335], '3-DehydroquinaseThe': [2337], 'aroD': [2338, 2442, 2492, 2527, 2615, 2630, 3099, 3823, 3892, 3925], 'gene': [2339, 2390, 2493, 2631, 3824], 'encoding': [2340], 'isolated,': [2349], 'free': [2350], 'extraneous': [2352, 3832], 'chromosomal': [2353, 3833, 3857], 'DNA,': [2354], 'plasmid': [2356, 2521], 'PCR': [2359, 2400, 3860], 'using': [2360, 2494, 3017, 3032, 3134, 3520, 3609, 3653, 3676, 3785], 'primers': [2361, 2372], 'complementary': [2362], 'ends': [2365], 'open': [2368], 'reading': [2369], 'frame.': [2370], 'extended': [2374], 'designed': [2378], 'place': [2380], 'EcoRI': [2382, 2451], 'restriction': [2383, 2448, 2485], '5′-end': [2387], 'HindIII': [2393, 2453], '3′-end.': [2397], 'blunt-ended': [2399], 'fragments': [2401], 'pUC18-derived': [2406], 'cloning': [2407, 3862], 'SmaI': [2412], 'overlapping': [2414], 'EcoK': [2416], 'polylinker,': [2420], 'introduced': [2422, 2465, 2524], 'JM103': [2426], 'EcoK+).': [2429], 'Clones': [2430], 'containing': [2431, 2476, 2980, 3717, 4302], 'inserts': [2432], 'selected': [2434, 3087], 'β-galactosidase': [2439], 'then': [2445, 2523, 2756, 2849, 2929, 3417], 'digestion': [2449], 'pKK223-3.': [2460], 'construct': [2463, 3846, 3881], 'JM105': [2469], 'transformation.': [2471], 'Plasmid': [2472], 'DNA': [2473, 2488, 2763, 3834, 3858], 'clones': [2475, 2501], 'insert': [2478], 'checked': [2480, 4168], 'appropriate': [2482], 'size': [2483, 2515, 3120], 'sites.': [2486], 'Following': [2487], 'A.L.F': [2496], 'automated': [2497], 'sequencer': [2498], '(Pharmacia),': [2499], 'suitable': [2500, 2514, 3922], 'induced': [2503, 2559], 'isopropyl': [2505, 2563], 'thiogalactoside': [2506], '(pAL2-2)': [2509], 'producing': [2510], 'band': [2512, 4214], 'SDS-PAGE': [2517, 2885, 4053, 4170], 'selected.': [2519], 'point': [2528, 3893], 'strain,': [2530], 'AB2848.': [2533], 'For': [2534, 2907, 3173, 3245, 3402, 3427, 3473, 3566], 'protein': [2535, 2846, 2966, 3248, 3391, 3438, 3481, 3561, 3579, 3755, 4151, 4206, 4343], 'preparation,': [2536], 'clone': [2538], 'grown': [2540, 2568, 2714, 3966, 4014], 'LB': [2542, 2716, 4016], 'medium': [2543, 2661, 2717, 3969, 4001, 4017], '200': [2548], 'μg/ml': [2549, 2603, 3252, 3485], 'ampicillin': [2550], 'OD550': [2553], '0.7-0.8,': [2555], 'whereupon': [2556], '1': [2561, 2728, 2830, 2833, 3216, 3828], 'mM': [2562, 2723, 2726, 2729, 2732, 2825, 2828, 2831, 2834, 2874, 2879, 2974, 2982, 3540, 4366, 4370], 'thiogalactoside.': [2564], '6': [2572, 4351], 'h': [2573, 3422], 'harvested': [2575], 'centrifugation': [2577, 2788, 2813], '5000': [2579], '×': [2580, 2791, 2815, 2942, 3218], 'g': [2581, 2792], '10': [2583, 3187], 'min.In': [2584], 'Vivo': [2585], 'Complementation': [2586], 'TestsStrain': [2587], 'inoculated': [2590], 'onto': [2591], 'M9': [2592], 'minimal': [2593, 3968, 4000], 'agar': [2594], 'plates': [2595], '(40': [2602], 'phenylalanine,': [2604], 'tyrosine,': [2605], 'tryptophan).': [2607], 'Bacteria': [2608], 'harboring': [2609], 'plasmids': [2611], 'enzymically': [2613], 'genes': [2616, 3103], '(wild': [2617], 'type,': [2618], 'M205L,': [2619], 'H146A)': [2621], 'grew': [2622, 2636, 2657], 'plates.': [2625], 'Those': [2626], '(pKK223-3': [2632], 'K170A)': [2635], 'supplemented': [2639, 2660], 'acids.': [2643], 'gave': [2647], 'rise': [2648], 'phenotype': [2652, 3960], 'bacteria': [2656], 'well': [2658, 4040], 'very': [2663], 'acids.Purification': [2668], 'DehydroquinasePurifications': [2670], 'overexpressed': [2672, 3109, 4039], 'carried': [2682, 2999, 3680], 'out': [2683, 3000, 3681], 'procedure': [2686], 'described': [2691, 3111, 3154, 3683, 3732, 3901, 4019, 4318], 'et': [2694, 3686, 4420], 'al.(10Chaudhuri': [2695], 'Cells': [2713], 'resuspended': [2719], 'buffer': [2721, 2823, 3448], '(50': [2722, 2824], 'Tris•HCl,': [2724, 2826, 3541], '5': [2725], 'MgCl2,': [2727], 'EDTA,': [2730, 2832], '1.2': [2731], 'phenylmethylsulfonyl': [2733], 'fluoride,': [2734], '8.0;': [2736], '40': [2737], 'ml/liter': [2738], 'culture': [2740], 'harvested)': [2741], 'treated': [2743], 'lysozyme': [2745], '(0.1': [2746], 'mg/ml)': [2747, 2781], '30': [2749, 2783, 2794, 2817, 3558, 3588, 4384, 4398], 'min': [2750, 2784, 2795], '37°C.': [2752, 2786], 'disrupted': [2757], 'three': [2759], 'freeze-thaw': [2760], 'cycles.': [2761], 'RNA': [2765], 'viscous': [2768], 'mixture': [2769, 3297, 3338, 3713, 4297], 'hydrolyzed': [2771], 'incubation': [2773], 'DNase': [2775], 'RNase': [2778], 'A': [2779, 2960, 3711], '(0.05': [2780], 'After': [2787], '21,000': [2790], '4°C,': [2797, 3416], 'supernatant': [2799], 'taken': [2801, 3514], '75%': [2803], 'saturation': [2804], 'ammonium': [2806], 'sulfate.': [2807], 'precipitate': [2809], 'recovered': [2811], '(21,000': [2814], 'g,': [2816], 'min),': [2818], 'redissolved': [2819], 'low': [2821], 'salt': [2822, 2871], '50': [2827, 2973, 2981, 4365, 4369], 'KCl,': [2829], '2-mercaptoethanol,': [2835], '7.5)': [2837], 'dialyzed': [2839, 2913, 3535], 'extensively': [2840], 'against': [2841, 2914, 2972, 3536], 'same': [2843, 2864, 2948], 'buffer.': [2844, 2865, 2949], 'solution': [2847], 'applied': [2850, 2930], '50-ml': [2853], 'column': [2854, 2937], 'anion': [2857], 'equilibrated': [2861, 2945], 'Dehydroquinase': [2866], 'eluted': [2868], 'gradient': [2872], '(50-250': [2873], 'KCl;': [2875], 'elution': [2876], '˜90': [2878], 'salt).': [2880], 'Fractions': [2881], 'assessed': [2883], 'usually': [2891], 'than': [2893, 3388, 3587], '85%': [2894], 'pure': [2895, 2957, 4176], 'stage.': [2898], 'Yields': [2899], 'up': [2901, 3489], '100': [2903], 'mg/liter': [2904, 3873], 'culture.': [2906], 'purification,': [2909, 4062], 'fractions': [2911], 'potassium': [2915, 2975, 3265], 'phosphate': [2916, 2976, 3266], 'buffer,': [2917, 2977, 3267], '7.0,': [2919, 2979, 3269], 'concentrated': [2920], '˜4': [2922], 'mg/ml': [2923], 'Amicon': [2926], 'concentrator': [2927], 'semipreparative': [2933], 'Superdex-S75': [2934], 'filtration': [2936], '(360': [2938], 'ml;': [2939], '26': [2940], 'mm': [2941], '600': [2943], 'mm)': [2944], 'Enzyme': [2950], 'way': [2953], 'typically': [2955], '>95%': [2956, 4175], 'SDS-PAGE.': [2959, 3126], 'dialyzing': [2968], 'purified': [2970, 4011], 'hydroxylamine': [2983], 'removed': [2985, 3455], 'base-linked': [2987], 'molecules': [2989, 3722], '(see': [2990, 3136, 3874], '“Isoelectric': [2991], 'Focusing”': [2992], 'under': [2993, 3617, 4020], '“Results”).Protein': [2994], 'DeterminationsProtein': [2995], 'concentration': [2996, 3373], 'determinations': [2997, 3191, 3626, 3782], 'Bradford(27Bradford': [3005], 'Anal.': [3007], '1976;': [3009], '72:': [3010], '248-254Crossref': [3011], '(213154)': [3014], 'kit': [3019], 'Bio-Rad,': [3021], 'bovine': [3023], 'serum': [3024], 'albumin': [3025], 'standard': [3028, 3632], 'spectrophotometry': [3031, 3149], 'extinction': [3034], 'coefficient': [3035], '∈280': [3036], '=': [3037], '19,000': [3038], '280': [3040], 'nm': [3041, 3152, 3234], '(19Deka': [3042], 'ScholarMutagenesisA': [3059], 'M13': [3061], 'generation': [3067], 'directed': [3070], 'mutants(28Curtis': [3071], 'M.D.': [3072], 'James': [3073], '5:': [3078], '2727-2733Crossref': [3079], '(29)': [3082], 'Mutants': [3085], 'track': [3090], 'confirmed': [3093, 3129], 'gene.': [3100], 'above.': [3112], 'similar': [3118, 4071, 4241], 'Mutations': [3127], 'accurate': [3131], 'measurement': [3133], 'ESI-MS': [3135], 'below).Enzyme': [3137], 'AssayDehydroquinase': [3138], 'assayed': [3141], 'monitoring': [3143], 'appearance': [3145], '234': [3151, 3364], 'previously(10Chaudhuri': [3155], 'determination': [3174, 3409, 3637], 'kinetic': [3176], 'parameters,': [3177], 'seven': [3178], 'concentrations': [3179, 3249, 3257, 3439, 3482, 3488, 4072], 'spanning': [3182], 'range': [3184, 3598], '0.1': [3185], 'Km': [3188, 3781], 'used;': [3190], 'done': [3193, 3509], 'separate': [3195, 3624], 'preparations': [3197], 'least': [3200, 3420], 'duplicate.SpectrofluorimetryExperiments': [3202], 'performed': [3204, 3784], 'Shimadzu': [3207], 'RF5000': [3208], 'spectrofluorimeter': [3209], 'interfaced': [3210], 'personal': [3213], 'computer,': [3214], 'cm': [3217], '1-cm': [3219], 'cuvettes': [3220], 'total': [3224], 'volume': [3225], '1.5': [3227], 'ml.': [3228], 'excitation': [3230], 'wavelength': [3231], '295': [3233], 'emission': [3236, 3272], 'spectra': [3237, 3445, 3512], 'acquired': [3239, 3595], '300': [3241], '450': [3243], 'nm.': [3244, 3277], 'Kd': [3246, 3408], 'determinations,': [3247], '˜50': [3251, 3484], '(˜2': [3253], 'μM),': [3254], 'ligand': [3256, 3286, 3453, 3470, 3487, 3774], '0': [3260], '160': [3262], 'μM,': [3263], '25°C;': [3271], 'monitored': [3274], '330': [3276], 'majority': [3289], 'equilibrium': [3296, 3305, 3358], 'dehydroquinate,': [3300], 'dehydroshikimate;': [3303], 'constant': [3306], 'Mitsuhashi': [3316], '(29Mitsuhashi': [3319], 'B.D.': [3322], 'Biochim.': [3323], 'Biophys.': [3324], 'Acta.': [3325], '1954;': [3326], '15:': [3327], '54-61Crossref': [3328], '(44)': [3331], '15.': [3336], 'generated': [3343], 'incubating': [3345], '(overnight': [3346], '4°C)': [3348], 'amounts': [3353], 'until': [3357], 'reached': [3361], '(monitored': [3362], 'nm)': [3365], 'treatment.': [3370], 'final': [3372], 'during': [3378, 4358], 'spectrofluorometric': [3380], 'assays': [3381], '˜3': [3383], 'orders': [3384], 'magnitude': [3386], 'lower': [3387, 3437], 'target': [3390], 'did': [3394, 3982], 'contribute': [3396], 'any': [3398, 4045, 4155, 4198], 'observed': [3399, 4046], 'changes.': [3401], 'samples': [3406], 'allowed': [3411], 'equilibrate': [3413], 'overnight': [3414], '2': [3421], '25°C': [3424], 'before': [3425, 3590], 'measurement.': [3426], 'necessary': [3433], 'use': [3435], 'much': [3436, 4003], '(1.25': [3440], 'μg/ml,': [3441], '0.05': [3442], 'μM).': [3443], 'Baseline': [3444], 'due': [3446, 4146], 'and,': [3449], 'where': [3450], 'employed,': [3451, 3494], 'subtraction': [3457], 'every': [3459], 'case,': [3460], 'fitted': [3466, 3500], 'curve.': [3472], 'time': [3474, 3568, 3777], '500': [3491], 'μM': [3492, 4382, 4396], 'exponential': [3504], 'decay.': [3505], 'All': [3506], 'duplicate.ESI-MSMass': [3511], 'Fisons': [3517], 'VG': [3518], 'Platform': [3519], '1:1:0.001': [3521], 'CH3CN:H2O:HCOOH': [3522], 'sample': [3524, 3551], 'solvent': [3525, 3552, 3572, 3584], 'horse': [3527], 'heart': [3528], 'myoglobin': [3529], 'standard.': [3532], 'Samples': [3533, 3750], '1-2': [3539], '7.5,': [3543], 'remove': [3545], 'salts,': [3546], 'mixed': [3548], 'immediately': [3553], 'prior': [3554], 'analysis.': [3556], 'Approximately': [3557], 'pmol': [3559], 'per': [3564, 3727], 'run.': [3565], 'experiments,': [3570, 3776, 3779], 'flow': [3573], 'rate': [3574], 'exposed': [3581], 'less': [3586], 's': [3589], 'ionization.': [3592], 'Data': [3593, 3791], 'over': [3596], 'm/z': [3599], '700-1300,': [3600], 'five': [3602, 3623], 'scans': [3605], 'averaged': [3606], 'processed': [3608], "supplier's": [3611], 'MassLynx': [3612], 'software.': [3613], 'Standard': [3614], 'errors': [3615, 3621], 'quoted': [3616], '“Results”': [3618], 'represent': [3619], 'sample.': [3629], 'typical': [3631], 'deviation': [3633], 'order': [3641], '1.0-2.0': [3643], 'Da.Isoelectric': [3644], 'Focusing': [3645], '(IEF)Samples': [3646], 'analyzed': [3648, 4189], 'PhastSystem': [3652], 'precast': [3654], 'IEF': [3655], 'gels': [3656, 3760, 4262], 'covering': [3657], 'pI': [3658, 4217], '4.0-6.5': [3659], 'standards': [3663], 'kit.': [3664], 'Irreversible': [3665], '(dehydroshikimate)': [3669], 'borohydride': [3678], 'al.(14Kleanthous': [3687, 4421], 'hybrid': [3712, 4296], 'composed': [3714], 'none,': [3718, 4306], 'one,': [3719, 4307], 'dimer': [3728], 'previously(30Reilly': [3733, 4319], '1429-1434Crossref': [3745, 4235, 4331], '(2)': [3747, 4237, 4333], '0.5-1': [3752], 'μg': [3753], 'loaded,': [3757], 'stained': [3762], 'Coomassie': [3764], 'Blue': [3765], 'according': [3766], "manufacturer's": [3769], 'protocol.Data': [3770], 'AnalysisCurve': [3771], 'fitting': [3772], 'Enzfitter': [3786], '(31Leatherbarrow': [3787], 'Enzfitter-Non-linear': [3789], 'Regression': [3790], 'Analysis': [3792], 'Program': [3793], 'IBM': [3796], 'PC.': [3797], 'Biosoft,': [3798, 3810], 'Cambridge,': [3799, 3811], 'UK1987Google': [3800], 'Ultrafit': [3803], '(32BiosoftUltrafit-Non-linear': [3804], 'Curve': [3805], 'Fitting': [3806], 'Package': [3807], 'Macintosh.': [3809], 'UK1992Google': [3812], 'software.RESULTSOverexpression': [3814], 'Purification': [3816], 'Dehydroquinases': [3818], 'AB2848pKD203': [3820], 'flanked': [3825], 'approximately': [3827], 'kilobase': [3829], 'pK223-3.': [3839], 'level': [3841, 3868], '˜10': [3848], 'mg/liter.': [3849], 'However,': [3850], 'removing': [3854], 'flanking': [3856], 'back': [3863], 'pK223-3': [3865], 'increased': [3866], '˜100': [3872], '“Materials': [3875, 4021], 'Methods”).': [3877], 'Using': [3878], 'transformed': [3889], 'AB2848,': [3895], 'Pittard': [3903], 'Wallace(26Pittard': [3905], 'chosen': [3919], 'deletion': [3926], 'unavailable.': [3929], 'auxotrophic': [3932], 'initial': [3943], 'assay': [3944], 'functional': [3946], 'Enzymatically': [3948], '(H146A': [3951], 'M205L)': [3953], 'complement': [3957], 'aroD-': [3959], 'inactive': [3979], 'not;': [3983], 'case': [3986], 'partial': [3992], 'complementation': [3993], 'seemed': [3994], 'occur': [3996], 'growth': [3998], 'slower.': [4004], 'Wild': [4005, 4208], 'Methods.”': [4023], 'physical': [4032], 'proteins;': [4036], 'toxic': [4042], 'degradation.': [4047], 'behaved': [4050], 'similarly': [4051], 'columns': [4059], 'preliminary': [4065], 'they': [4067], 'denatured': [4069], 'guanidine': [4074, 4353], 'those': [4077], 'reported': [4078], 'enzyme(3Kleanthous': [4084], '14Kleanthous': [4112], 'therefore': [4138], 'unlikely': [4139], 'specific': [4142], 'seen': [4144], 'destabilization': [4148], 'structure': [4152], 'caused': [4153], 'mutations.Isoelectric': [4158], 'FocusingThe': [4159], 'purity': [4160], '(data': [4177], 'shown).': [4179], 'matter': [4182], 'routine,': [4184], 'isoelectric': [4191, 4242], 'focusing': [4192, 4261], 'electrophoresis': [4194], 'whether': [4197], 'variants': [4200], 'apparent': [4202], 'native': [4205], 'preparations.': [4207], '4.9': [4218], '(in': [4219], 'agreement': [4220], 'previous': [4222], 'work)(30Reilly': [4223], 'points': [4243], 'M205L': [4250, 4289], '1A,': [4253], 'lanes': [4254], '1,': [4255, 4271, 4342, 4402], '3,': [4256, 4282, 4361, 4446], '4,': [4258, 4288, 4376, 4453], 'respectively).Figure': [4259], '1Isoelectric': [4260], 'dehydroquinases.': [4268], 'A,': [4269], 'lane': [4270, 4275, 4281, 4287, 4293, 4346, 4360, 4375, 4386, 4401, 4407, 4445, 4452], 'DHQase;': [4274], '2,': [4276, 4347, 4408], 'prepared;': [4280, 4286, 4292, 4345, 4406, 4451], '5,': [4294, 4387], 'subunits': [4310], 'labeled': [4311], 'dehydroshikimate,': [4315], 'B,': [4336], 'treatments': [4337], 'Lane': [4341], 'after': [4348, 4362, 4377, 4391, 4411, 4457], 'unfolding': [4349], 'M': [4352], 'followed': [4355], 'refolding': [4357], 'dialysis;': [4359], 'dialysis': [4363], 'NH2OH•HCl': [4367], '+': [4368], 'Tris,': [4371], '7.0': [4373], '(final);': [4374], '(500': [4381, 4395], 'min);': [4385], 'min).': [4399], 'C,': [4400], 'NaBH4': [4416], '(according': [4417], 'Scholar));': [4444], 'NaBH4(14Kleanthous': [4462], 'Scholar).View': [4485], 'Large': [4486], 'Image': [4487], 'Figure': [4488], 'ViewerDownload': [4489], 'Hi-res': [4490]}, 'cited_by_api_url': 'https://api.openalex.org/works?filter=cites:W1993808586', 'counts_by_year': [{'year': 2024, 'cited_by_count': 1}, {'year': 2023, 'cited_by_count': 5}, {'year': 2022, 'cited_by_count': 2}, {'year': 2021, 'cited_by_count': 1}, {'year': 2020, 'cited_by_count': 2}, {'year': 2015, 'cited_by_count': 4}, {'year': 2014, 'cited_by_count': 1}, {'year': 2013, 'cited_by_count': 2}, {'year': 2012, 'cited_by_count': 3}], 'updated_date': '2024-12-11T15:27:02.855380', 'created_date': '2016-06-24'}
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