[Molecular form and stability of D-hydantoinase deleted at C-terminal residue Arg].
This report is about only deleting one C-terminal residue of D-hydantoinase to result in obvious changes on its molecular form and stability. A recombinant D-hydantoinase (P479) and its mutant enzyme deleted at C-terminal residue Arg (P478) were prepared by methods of gene cloning, expression and purification. Results show that the subunit molecular weight of P479 and P478 is the same (54kDa) as determined by SDS-PAGE, whilst the molecular form of native P479 and P478 is a dimer and a monomer respectively in the completely operative conditions. Compared with P479, the enzymatic activity of P478 for substrate hydantoin maintained about 40% and pH stability was obviously increased, at the alkaline side in particular, as well as the anti-SDS ability was also raised. However, the thermal stability for P478 was clearly lowed as compared to P479. It implies from above data that the C-terminal residue Arg of the D-hydantoinase is a crucial one for subunit dissociation, but non-essential for catalysis.
Niu
Li-xi
LX
Institute of Biotechnology, Key Laboratory of Chemical Biology and Molecular Engineering of National Ministry of Education, Shanxi University, Taiyuan 030006, China.
Zhang
Xue-yao
XY
Shi
Ya-wei
YW
Yuan
Jing-ming
JM
D000581:Amidohydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D009154:Mutation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology; D013696:Temperature
D011994:Recombinant Proteins; D012967:Sodium Dodecyl Sulfate; D000581:Amidohydrolases; C020047:dihydropyrimidinase