Role of conserved histidine residues in D-aminoacylase from Alcaligenes xylosoxydans subsp. xylosoxydans A-6

Article abstract of DOI:10.1271/bbb.64.1

D-Aminoacylase from Alcaligenes xylosoxydans subsp. xylosoxydans A-6 (Alcaligenes A-6) was strongly inactivated by diethylpyrocarbonate (DEPC). An H67N mutant was barely active, with a k_cat/K_m 6.3 × 10⁴ times lower than that of the recombinant wild-type enzyme, while the H67I mutant lost detectable activity. The H67N mutant had almost constant K_m, but greatly decreased k_cat. These results suggested that His67 is essential to the catalytic event. Both H69N and H69I mutants were overproduced in the insoluble fraction. The k_cat/K_m of H250N mutant was reduced by a factor of 2.5 × 10⁴-old as compared with the wild-type enzyme. No significant difference between H251N mutant and wild-type enzymes in the K_m and k_cat was found. The Zn content of H250N mutant was nearly half of that of wild-type enzyme. These results suggest that the His250 residue might be essential to catalysis via Zn binding.

Full text of DOI:10.1271/bbb.64.1

Bioscience, Biotechnology, and Biochemistry
ISSN: 0916-8451 (Print) 1347-6947 (Online) Journal homepage: http://www.tandfonline.com/loi/tbbb20
Role of Conserved Histidine Residues in D-
Aminoacylase from Alcaligenes xylosoxydans
subsp. xylosoxydans A-6
Mamoru WAKAYAMA, Harutaka YADA, Shun-ichi KANDA, Shin-ichi HAYASHI,
Yukinori YATSUDA, Kenji SAKAI & Mitsuaki MORIGUCHI
To cite this article: Mamoru WAKAYAMA, Harutaka YADA, Shun-ichi KANDA, Shin-ichi HAYASHI,
Yukinori YATSUDA, Kenji SAKAI & Mitsuaki MORIGUCHI (2000) Role of Conserved Histidine
Residues in D-Aminoacylase from Alcaligenes xylosoxydans subsp. xylosoxydans A-6,
Bioscience, Biotechnology, and Biochemistry, 64:1, 1-8, DOI: 10.1271/bbb.64.1
To link to this article: http://dx.doi.org/10.1271/bbb.64.1
Published online: 22 May 2014.
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Date: 04 June 2016, At: 23:26