Mechanism of Proteinase Release from Lactococcus lactis subsp. cremoris Wg2
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The procedure generally used for the isolation of extracellular, cell-associated proteinases of Lactococcus lactis species is based on the release of the proteinases by repeated incubation and washing of the cells in a Ca-free buffer. For L. lactis subsp. cremoris Wg2, as many as five incubations for 30 min at 29 degrees C are needed in order to liberate 95% of the proteinase. Proteinase release was not affected by chloramphenicol, which indicates that release is not the result of protein synthesis during the incubations. Ca inhibited, while ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid (EGTA) stimulated, proteinase release from the cells. The pH optimum for proteinase release ranged between 6.5 and 7.5, which was higher than the optimum pH of the proteinase measured for casein hydrolysis (i.e., 6.4). Treatment of cells with the serine proteinase inhibitor phenylmethylsulfonyl fluoride prior to the incubations in Ca-free buffer reduced the release of the proteinase by 70 to 80%. The residual proteinase remained cell associated but could be removed by the addition of active L. lactis subsp. cremoris Wg2 proteinase. This suggests that proteinase release from cells of L. lactis subsp. cremoris Wg2 is the result of autoproteolytic activity. From a comparison of the N-terminal amino acid sequence of the released proteinase with the complete amino acid sequence determined from the nucleotide sequence of the proteinase gene, a protein of 180 kilodaltons would be expected. However, a proteinase with a molecular weight of 165,000 was found, which indicated that further hydrolysis had occurred at the C terminus.
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The residual PrtP activity after 24 h was decreased to about 0.1% of the initial activity.
“The residual PrtP activity was decreased to about 0.1% of the initial activity.”
From The extracellular PI-type proteinase of Lactococcus lactis hydrolyzes beta-casein into more than one hundred different oligopeptides (1995) · cites this paper as [18]
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- Authors
- Laan, H., & Konings, W. N.
- Affiliations
- University of Groningen (NL)
- Venue
- Applied and Environmental Microbiology
- Publisher
- American Society for Microbiology
- Published
- 1989-12
- Volume
- 55
- Issue
- 12
- Pages
- 3101-3106
- ISSN
- 0099-2240 (ISSN-L)
- 1098-5336
- PMC
- PMC203230
- OpenAlex
- W1590961427
- Cited by
- 76
- References
- 28
- Citations by year
- 2025: 1 · 2022: 1 · 2021: 1 · 2020: 1 · 2019: 2 · 2018: 1 · 2017: 1 · 2015: 1 · 2014: 1 · 2013: 1 · 2012: 4
- FWCI
- 11.63
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Locations
- Applied and Environmental Microbiology (publishedVersion)
- PubMed Central (publishedVersion)
- PubMed (publishedVersion)
- University of Groningen research database (University of Groningen / Centre for Information Technology) (submittedVersion)
- Topics
- Life Sciences › Agricultural and Biological Sciences › Food Science › Probiotics and Fermented Foods
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Biotechnology › Enzyme Production and Characterization
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Bacterial Genetics and Biotechnology
- Keywords
- Lactococcus lactis subsp. cremoris, extracellular protease, calcium dependence, EGTA, phenylmethylsulfonyl fluoride, serine protease, casein hydrolysis, N-terminal sequence, cell-associated enzymes
About these checks
Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.
Statistics and arithmetic are recomputed, not judged.
Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.
What counts as a claim ›How we measure confidence ›How we check this ›
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