Lactococcal proteinase maturation protein PrtM is a lipoprotein.
The production of enzymatically active proteinase by lactococci requires the joint presence of a proteinase gene, prtP, and a gene encoding a maturation protein, prtM. A 32-kDa protein produced by Escherichia coli upon expression of the prtM gene under the direction of the T7 RNA polymerase promoter was purified and used to obtain PrtM-specific antibodies. With these antibodies, immunogold labeling of lactococcal cells revealed that PrtM was associated with the lactococcal cell envelope. Western blot (immunoblot) analysis of whole lactococcal cells and isolated membrane vesicles indicated that PrtM was a membrane-associated protein. Radiolabeling of Lactococcus lactis with [3H]palmitic acid showed that PrtM was a lipoprotein. Partial secretion of PrtM into the culture medium was observed after Cys-24, the target residue for lipid modification, was replaced by an Ala residue by means of site-directed mutagenesis. This mutation did not affect proteinase activity.
Haandrikman
A J
AJ
Department of Genetics, University of Groningen, Haren, The Netherlands.
Kok
J
J
Venema
G
G
D000595:Amino Acid Sequence; D001483:Base Sequence; D015153:Blotting, Western; D003001:Cloning, Molecular; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D005838:Genotype; D013294:Lactococcus lactis / Q000235:genetics*; D008074:Lipoproteins / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D008894:Milk Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D019308:Palmitic Acid; D010169:Palmitic Acids / Q000378:metabolism; D010957:Plasmids; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid
D008074:Lipoproteins; D008894:Milk Proteins; D010169:Palmitic Acids; D011994:Recombinant Proteins; D019308:Palmitic Acid; D010450:Endopeptidases