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J Bacteriol. 1999 Oct;181(19):6033-41.

A mutant HemA protein with positive charge close to the N terminus is stabilized against heme-regulated proteolysis in Salmonella typhimurium.

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  • 1Department of Microbiology and Immunology, West Virginia University Health Sciences Center, Morgantown, West Virginia 26506, USA.

Abstract

The HemA enzyme (glutamyl-tRNA reductase) catalyzes the first committed step in heme biosynthesis in the enteric bacteria. HemA is mainly regulated by conditional protein stability; it is stable and, consequently, more abundant in heme-limited cells but unstable and less abundant in normally growing cells. Both the Lon and ClpAP energy-dependent proteases contribute to HemA turnover in vivo. Here we report that the addition of two positively charged lysine residues to the third and fourth positions at the HemA N terminus resulted in complete stabilization of the protein. By contrast, the addition of an N-terminal myc epitope tag did not affect turnover. This result confirms the importance of the N-terminal sequence for proteolysis of HemA. This region of the protein also contains a proline flanked by hydrophobic residues, a motif that has been suggested to be important for Lon-mediated proteolysis of UmuD. However, mutation of this motif did not affect the turnover of HemA protein. Cells expressing the stabilized HemA[KK] mutant protein display substantial defects in heme regulation.

PMID:
10498716
[PubMed - indexed for MEDLINE]
PMCID:
PMC103631
Free PMC Article
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