RRC ID 55093
著者 Pei X, Wang J, Wu Y, Zhen X, Tang M, Wang Q, Wang A.
タイトル Evidence for the participation of an extra α-helix at β-subunit surface in the thermal stability of Co-type nitrile hydratase.
ジャーナル Appl Microbiol Biotechnol
Abstract Nitrile hydratase (NHase) has attracted considerable attention since it can efficiently catalyze the hydration of nitriles to valuable amides. However, the poor stability of NHase is one of the main drawbacks in the industrial application. In this study, we compared the structural difference between Fe-type and Co-type NHase and found that an extra α helix existed at the β-subunit surface of Co-type NHase (defined as the β-6th helix). Then, the effects of the β-6th helix were investigated on the thermal stability and the catalytic kinetics of a Co-type NHase from Aurantimonas manganoxydans ATCC BAA-1229 (NHase1229). When the β-6th helix was deleted or disrupted, the thermal stability of NHase1229 was reduced to 17.6 and 12.9% of that of wild NHase1229, respectively. Thus, the β-6th helix is important for the thermal stability of Co-type NHase. Based on the structural characteristics of Co-type NHase, the β-6th helix may be interacted with another helix at the α-subunit (defined as the α-2nd helix) by hydrophobic network just as a "magnetic suction buckle" on the enzyme surface to stabilize the binding of α- and β-subunits. The β-6th helix is located at the mouth of the substrate and product tunnel, so it plays crucial roles in catalytic process. Furthermore, the β-6th helix in NHase1229 was swapped with a thermophilic NHase fragment from Pseudonocardia thermophila JCM3095 (NHase1229-Swap). The thermal stability of NHase1229-Swap was significantly improved, and the half-life was approximately 2.4-fold at 40 °C than that of the wild NHase1229. The knowledge is useful for improving the stability of NHases by restriction fragment swapping.
巻・号 102(18)
ページ 7891-7900
公開日 2018-9-1
DOI 10.1007/s00253-018-9191-2
PII 10.1007/s00253-018-9191-2
PMID 29998413
MeSH Amino Acid Sequence Cobalt / chemistry* Enzyme Stability Hot Temperature* Hydro-Lyases / chemistry* Hydro-Lyases / metabolism* Hydrophobic and Hydrophilic Interactions
IF 3.67
引用数 4
リソース情報
一般微生物 JCM 3095