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https://doi.org/10.5562/cca2107

Yeast Ortholog of Peptidase Family M49: the Role of Invariant Glu461 and Tyr327

Nina Jajčanin-Jozić ; Division of Organic Chemistry and Biochemistry, Ruđer Bošković Institute, Bijenička cesta 54, HR-10002 Zagreb, Croatia
Peter Macheroux ; Institute of Biochemistry, Graz University of Technology, Petersgasse 12, A-8010 Graz, Austria
Marija Abramić ; Division of Organic Chemistry and Biochemistry, Ruđer Bošković Institute, Bijenička cesta 54, HR-10002 Zagreb, Croatia


Puni tekst: engleski pdf 2.635 Kb

str. 535-540

preuzimanja: 983

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Sažetak

Metallopeptidase family M49 is characterized by five conserved sequence regions and the unique motif HEXXGH with two histidines - ligands of the active-site zinc ion. The crystal structure of the yeast ortholog represents a prototype for the whole family.
To investigate the role of two invariant amino acid residues, a Glu461 of the zinc-binding motif, and a Tyr327, 21Å from the catalytic zinc center, mutational analysis of the yeast enzyme was performed.
The substitution of Glu461 to glutamine decreased kcat for the substrate hydrolysis almost by 10 000-fold. The replacement of Tyr327 by Phe or Ala reduced the catalytic efficiency (kcat/Km) by two orders of magnitude. The affinity for the heptapeptide valorphin was siginificantly lowered in all mutants, in-dicating the contribution of both Glu461 and Tyr327 in substrate binding. Taken together, the effect of mutating Glu461 is consistent with this residue being essential in M49 peptidase catalysis. (doi: 10.5562/cca2107)

Ključne riječi

enzyme catalysis, metallopeptidase, protein structure-function, site-directed mutagenesis, yeast Saccharomyces cerevisiae

Hrčak ID:

94463

URI

https://hrcak.srce.hr/94463

Posjeta: 1.544 *