Biotechnology Bulletin ›› 2021, Vol. 37 ›› Issue (3): 75-83.doi: 10.13560/j.cnki.biotech.bull.1985.2020-0830
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HAO Jun-yao1(), MA Fu-qiang1,2, YANG Guang-yu1
Received:
2020-07-07
Online:
2021-03-26
Published:
2021-04-02
HAO Jun-yao, MA Fu-qiang, YANG Guang-yu. Functional Analysis of Key Residues in the Active Center of Creatinase from Alcaligenes sp. KS-85[J]. Biotechnology Bulletin, 2021, 37(3): 75-83.
氨基酸残基 | 位置 | 功能 |
---|---|---|
Arg66 | loop | 底物结合 |
Ile84 | 310 helix | 稳定过渡态 |
His233 | loop | 酸碱催化 |
Tyr259 | β sheet | 稳定过渡态 |
Glu263 | β sheet | 底物结合 |
His325 | loop | 稳定过渡态 |
Arg336 | loop | 底物结合 |
Glu359 | β sheet | 稳定过渡态 |
His377 | β sheet | 稳定过渡态 |
氨基酸残基 | 位置 | 功能 |
---|---|---|
Arg66 | loop | 底物结合 |
Ile84 | 310 helix | 稳定过渡态 |
His233 | loop | 酸碱催化 |
Tyr259 | β sheet | 稳定过渡态 |
Glu263 | β sheet | 底物结合 |
His325 | loop | 稳定过渡态 |
Arg336 | loop | 底物结合 |
Glu359 | β sheet | 稳定过渡态 |
His377 | β sheet | 稳定过渡态 |
突变体 | kcat(min-1) | KM(mM) | kcat/KM(mM-1·min-1) |
---|---|---|---|
WT | 108.7±1.43 | 2.12±0.33 | 51.27 |
F64A | 13.603±2.06 | 125.00±3.02a | 0.11 |
F64S | 30.66±1.66 | 282.20±19.61a | 0.11 |
F64D | ND | ND | ND |
F64R | 17.91±1.04 | 98.57±11.21a | 0.18 |
F64Y | 28.47±1.38 | 10.15±1.85 | 2.80 |
F64W | 37.84±0.68 | 13.82±0.97 | 2.73 |
D102A | 8.28±0.52 | 29.1±2.69 | 0.28 |
D102S | 1.056±0.07 | 51.64±3.56 | 0.05 |
D102R | 1.60±0.12 | 27.37±8.26 | 0.05 |
D102F | 4.97±0.04 | 37.85±14.77 | 0.13 |
D102Y | ND | ND | ND |
D102W | 13.13±0.10 | 25.78±0.94 | 0.50 |
F252A | 1.60±0.037 | 1.36±0.099 | 1.18 |
F252S | 2.50±0.033 | 0.46±0.03 | 5.47 |
F252D | ND | ND | ND |
F252R | ND | ND | ND |
F252Y | 11.92±0.12 | 0.836±0.01 | 14.25 |
F252W | 0.402±0.03 | 0.931±0.22 | 0.43 |
F321A | 1.34±0.09 | 1.72±0.15 | 0.77 |
F321S | 2.42±0.02 | 6.68±0.27 | 0.36 |
F321D | 0.54±0.01 | 20.64±1.30 | 0.03 |
F321R | 0.62±0.97 | 3.959±0.52 | 0.16 |
F321Y | 2.15±0.02 | 0.51±0.05 | 4.25 |
F321W | 0.32±0.01 | 23.35±2.09 | 0.01 |
突变体 | kcat(min-1) | KM(mM) | kcat/KM(mM-1·min-1) |
---|---|---|---|
WT | 108.7±1.43 | 2.12±0.33 | 51.27 |
F64A | 13.603±2.06 | 125.00±3.02a | 0.11 |
F64S | 30.66±1.66 | 282.20±19.61a | 0.11 |
F64D | ND | ND | ND |
F64R | 17.91±1.04 | 98.57±11.21a | 0.18 |
F64Y | 28.47±1.38 | 10.15±1.85 | 2.80 |
F64W | 37.84±0.68 | 13.82±0.97 | 2.73 |
D102A | 8.28±0.52 | 29.1±2.69 | 0.28 |
D102S | 1.056±0.07 | 51.64±3.56 | 0.05 |
D102R | 1.60±0.12 | 27.37±8.26 | 0.05 |
D102F | 4.97±0.04 | 37.85±14.77 | 0.13 |
D102Y | ND | ND | ND |
D102W | 13.13±0.10 | 25.78±0.94 | 0.50 |
F252A | 1.60±0.037 | 1.36±0.099 | 1.18 |
F252S | 2.50±0.033 | 0.46±0.03 | 5.47 |
F252D | ND | ND | ND |
F252R | ND | ND | ND |
F252Y | 11.92±0.12 | 0.836±0.01 | 14.25 |
F252W | 0.402±0.03 | 0.931±0.22 | 0.43 |
F321A | 1.34±0.09 | 1.72±0.15 | 0.77 |
F321S | 2.42±0.02 | 6.68±0.27 | 0.36 |
F321D | 0.54±0.01 | 20.64±1.30 | 0.03 |
F321R | 0.62±0.97 | 3.959±0.52 | 0.16 |
F321Y | 2.15±0.02 | 0.51±0.05 | 4.25 |
F321W | 0.32±0.01 | 23.35±2.09 | 0.01 |
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