Cloning, sequence, and phenotypic expression of katA, which encodes the catalase of Lactobacillus sake LTH677

H. J. Knauf, R. F. Vogel, W. P. Hammes

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Abstract

Lactobacillus sake LTH677 is a strain, isolated from fermented sausage, which forms a heme-dependent catalase. This rare property is highly desirable in sausage fermentation, as it prevents rancidity and discoloration caused by hydrogen peroxide. A gene bank containing MboI fragments of chromosomal DNA from Lactobacillus sake LTH677 in Escherichia coli plasmid pBR328 was constructed. The catalase gene was cloned by heterologous complementation of the Kat- phenotype of E. coli UM2. The catalase structural gene, designated katA, was assigned to a 2.3-kb region by deletion analysis of the originally cloned fragment in plasmid pHK1000. The original chromosomal arrangement was determined by Southern hybridization. Protein analysis revealed that the catalase subunit has a molecular size of 65,000 Da and that the active catalase possesses a hexameric structure. The molecular size of the subunit deduced from the nucleotide sequence was determined to 54,504 Da. The N- terminal amino acid sequence of the 65,000-Da protein corresponded to the one deduced from the DNA sequence. After recloning of katA in the E. coli- Lactococcus shuttle vector pGKV210, the gene was successfully transferred and phenotypically expressed in Lactobacillus casei, which is naturally deficient in catalase activity.

Original languageEnglish
Pages (from-to)832-839
Number of pages8
JournalApplied and Environmental Microbiology
Volume58
Issue number3
DOIs
StatePublished - 1992
Externally publishedYes

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