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Biochemical and Mutational Analysis of a Novel Nicotinamidase from Oceanobacillus iheyensis HTE831.
- Source :
- PLoS ONE; Feb2013, Vol. 8 Issue 2, p1-14, 14p
- Publication Year :
- 2013
-
Abstract
- Nicotinamidases catalyze the hydrolysis of nicotinamide to nicotinic acid and ammonia, an important reaction in the NAD<superscript>+</superscript> salvage pathway. This paper reports a new nicotinamidase from the deep-sea extremely halotolerant and alkaliphilic Oceanobacillus iheyensis HTE831 (OiNIC). The enzyme was active towards nicotinamide and several analogues, including the prodrug pyrazinamide. The enzyme was more nicotinamidase (k<subscript>cat</subscript>/K<subscript>m</subscript> = 43.5 mM<superscript>−1</superscript>s<superscript>−1</superscript>) than pyrazinamidase (k<subscript>cat</subscript>/K<subscript>m</subscript> = 3.2 mM<superscript>−1</superscript>s<superscript>−1</superscript>). Mutational analysis was carried out on seven critical amino acids, confirming for the first time the importance of Cys133 and Phe68 residues for increasing pyrazinamidase activity 2.9- and 2.5-fold, respectively. In addition, the change in the fourth residue involved in the ion metal binding (Glu65) was detrimental to pyrazinamidase activity, decreasing it 6-fold. This residue was also involved in a new distinct structural motif DAHXXXDXXHPE described in this paper for Firmicutes nicotinamidases. Phylogenetic analysis revealed that OiNIC is the first nicotinamidase described for the order Bacillales. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 19326203
- Volume :
- 8
- Issue :
- 2
- Database :
- Complementary Index
- Journal :
- PLoS ONE
- Publication Type :
- Academic Journal
- Accession number :
- 87624871
- Full Text :
- https://doi.org/10.1371/journal.pone.0056727