[Improving the activity of creatinase from Alcaligenes sp. KS-85 through semi-rational design].
Sheng Wu Gong Cheng Xue Bao
; 38(12): 4601-4614, 2022 Dec 25.
Article
en Zh
| MEDLINE
| ID: mdl-36593196
ABSTRACT
Creatinine levels in biological fluids are important indicators for the clinical evaluation of renal function. Creatinase (CRE, EC3.5.3.3) is one of the key enzymes in the enzymatic measurement of creatinine concentration, and it is also the rate-limiting enzyme in the whole enzymatic cascade system. The poor catalytic activity of CRE severely limits its clinical and industrial applications. To address this issue, a semi-rational design is applied to increase the activity of a creatinase from Alcaligenes sp. KS-85 (Al-CRE). By high-throughput screen of saturation mutagenesis libraries on the selected hotspot mutations, multiple variant enzymes with increased activity are obtained. The five-point best variant enzyme (I304L/F395V/K351V/Y63S/Q88A) were further obtained by recombine the improved mutations sites that to showed a 2.18-fold increased specific activity. Additionally, structure analysis is conducted to understand the mechanism of the activity change. This study paves the way for a better practical application of creatinase and may help further understand its catalytic mechanism.
Palabras clave
Texto completo:
1
Base de datos:
MEDLINE
Asunto principal:
Ureohidrolasas
Idioma:
Zh
Revista:
Sheng Wu Gong Cheng Xue Bao
Asunto de la revista:
BIOTECNOLOGIA
Año:
2022
Tipo del documento:
Article
País de afiliación:
China