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PMID: 8526515 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

Genetic and biochemical characterization of the Lactobacillus delbrueckii subsp. lactis bacteriophage LL-H lysin.

Applied and environmental microbiology ·Vol. 61 ·No. 11 ·1995-11-00 ·Pages 4004-11

Vasala A, Välkkilä M, Caldentey J, Alatossava T

Abstract

LL-H, a virulent phage of Lactobacillus delbrueckii subsp. lactis, produces a peptidoglycan-degrading enzyme, Mur, that is effective on L. delbrueckii, Lactobacillus acidophilus, Lactobacillus helveticus, and Pediococcus damnosus cell walls. In this study, the LL-H gene mur was cloned into Escherichia coli, its nucleotide sequence was determined, and the enzyme produced in E. coli was purified and biochemically characterized. Mur was purified 112-fold by means of ammonium sulfate precipitation and cation-exchange chromatography. The cell wall-hydrolyzing activity was found to be associated with a 34-kDa protein. The C-terminal domain of Mur is not essential for catalytic activity since it can be removed without destroying the lytic activity. The N-terminal sequence of the purified lysin was identical to that deduced from the nucleotide sequence, but the first methionine is absent from the mature protein. The N-terminal part of this 297-amino-acid protein had homology with several Chalaropsis-type lysozymes. Reduction of purified and Mur-digested L. delbrueckii cell wall material with labeled NaB3H4 indicated that the enzyme is a muramidase. The temperature optimum of purified Mur is between 30 and 40 degrees C, and the pH optimum is around 5.0. The LL-H lysin Mur is stable at temperatures below 60 degrees C.

MeSH Terms
Amino Acid Sequence Bacteriophages/enzymology,genetics,pathogenicity Base Sequence Cell Wall/metabolism Cloning, Molecular DNA Primers/genetics DNA, Viral/genetics Enzyme Stability Escherichia coli/genetics Gene Deletion Genes, Viral Lactobacillus/virology Molecular Sequence Data Muramidase/genetics,isolation & purification,metabolism Sequence Homology, Amino Acid Species Specificity Viral Proteins/genetics,isolation & purification,metabolism
Chemicals
DNA Primers DNA, Viral Viral Proteins Muramidase
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Vasala A
Department of Genetics, University of Oulu, Finland.
Välkkilä M
Caldentey J
Alatossava T
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Article Info
Journal
Applied and environmental microbiology
Abbr.
Appl Environ Microbiol
ISSN
0099-2240
DOI
10.1128/aem.61.11.4004-4011.1995
Published
1995-11-00
Pages
4004-11
Language
English
Region
United States
NLM ID
7605801
PMCID
PMC167708
Subset
IM
Databases
GENBANK
L42315
External Links
PubMed source DOI: 10.1128/aem.61.11.4004-4011.1995 PMC Full Text View journal details
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