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. 1999 Aug;181(15):4592-7.

Genetic Characterization of a Cell Envelope-Associated Proteinase From Lactobacillus Helveticus CNRZ32

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Free PMC article

Genetic Characterization of a Cell Envelope-Associated Proteinase From Lactobacillus Helveticus CNRZ32

J A Pederson et al. J Bacteriol. .
Free PMC article

Abstract

A cell envelope-associated proteinase gene (prtH) was identified in Lactobacillus helveticus CNRZ32. The prtH gene encodes a protein of 1,849 amino acids and with a predicted molecular mass of 204 kDa. The deduced amino acid sequence of the prtH product has significant identity (45%) to that of the lactococcal PrtP proteinases. Southern blot analysis indicates that prtH is not broadly distributed within L. helveticus. A prtH deletion mutant of CNRZ32 was constructed to evaluate the physiological role of PrtH. PrtH is not required for rapid growth or fast acid production in milk by CNRZ32. Cell surface proteinase activity and specificity were determined by hydrolysis of alpha(s1)-casein fragment 1-23 by whole cells. A comparison of CNRZ32 and its prtH deletion mutant indicates that CNRZ32 has at least two cell surface proteinases that differ in substrate specificity.

Figures

FIG. 1
FIG. 1
Partial nucleotide and deduced amino acid sequence of the L. helveticus CNRZ32 prtH gene. The arrowhead indicates the start of transcription. A putative Shine-Delgarno sequence is underlined. The putative cleavage site for the pre-pro-region is marked with a vertical arrow. The putative active-site residues are boxed. Long horizontal arrows indicate primers used for DNA probe synthesis. The stop codon is indicated with an asterisk. Short horizontal arrows indicate the putative transcriptional terminator.
FIG. 2
FIG. 2
Reverse-phase HPLC patterns of the hydrolysis products from αs1-CN (f1-23) after 15 min of incubation with wild-type CNRZ32 whole cells (A) and CNRZ32 prtH deletion mutant whole cells (B).

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