Proteolytic enzymes of dairy starter cultures

Thomas, T. D.; Pritchard, G. G.

doi:10.1111/j.1574-6968.1987.tb02464.x

Cited by 306 publications

(167 citation statements)

References 88 publications

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“…Degradation of milk proteins is essential for growth of these bacteria on milk products as well as for the development of the products' texture and flavour (Fox, 1989;Law & Kolstad, 1983 ; Thomas & Mills, 1981). Thus, knowledge as to the types of proteinases that appear extracellularly during growth of lactococci -the substrate specificity and biochemical characteristics of the proteinases, and how the enzymes vary in different strains -is of importance in the use of lactococci in food production.…”

Section: Introductionmentioning

confidence: 99%

Purification and characterization of the free form of the lactococcal extracellular proteinase and its autoproteolytic cleavage products

Nissen‐Meyer¹,

Sletten²

1991

Journal of General Microbiology

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In addition to the cell wall proteinase, Lactococcus lactis subsp. cremoris produced significant amounts of a free extracellular proteinase. The free proteinase activity was highest in the late exponential and early stationary phase of growth, whereas the cell wall activity was highest in the last half of the exponential phase. Both proteinase forms had a pH optimum between 4-6 and 5.8, and they behaved similarly upon anion exchange and hydrophobic interaction chromatography, chromatofocusing and gel filtration, indicating that they were related. Purification to homogeneity, as judged by SDS-PAGE, resulted in a 50O00-80000-fold increase in the specific activity of the free proteinase. It contained two major protein species (termed pro150 and proll5) with proteinase activity. As judged by SDS-PAGE, the M, values of pro150 and pro115 were 150000 and 115000, respectively, and by chromatofocusing the isoelectric points were 4.3 and 4.1, respectively. Upon gel filtration, pro150 and pro115 had M, values of 300000 and 125000, respectively, indicating that pro150 was a dimer and pro115 a monomer. Pro115 was an autodegradation product of prol50. Other distinct autodegradation products had M, values of 90000 (p90), 53000 (p53), 37000 (p37) and 30000 (p30). These had little if any proteinase activity. Proll5, p90 and p53 had a common N-terminal sequence with that reported for the cell wall proteinase. Judging from its N-terminal sequence and M,, p30 was derived from the C-terminal half of p53. Cleavage of pro150 to pro115 generated p37.

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Section: Introductionmentioning

confidence: 99%

Purification and characterization of the free form of the lactococcal extracellular proteinase and its autoproteolytic cleavage products

Nissen‐Meyer¹,

Sletten²

1991

Journal of General Microbiology

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“…Due to the rather low content of free essential amino acids in milk they are dependent on their proteolytic abilities to degrade milk casein. Lactobacilli possess a complex proteolytic system composed of proteinases and a variety of peptidases (Abo-Elnaga & Plapp, 1987;Law & Kolstadt, 1983;Thomas & Pritchard, 1987), which act in a cascade to hydrolyse casein to small transportable peptides and amino acids. The quality of dairy products The GenBanWEMBUDDBJ accession number for the nucleotide sequence reported in this paper is 231377.…”

Section: Introductionmentioning

confidence: 99%

Cloning and nucleotide sequence analysis of pepV, a carnosinase gene from Lactobacillus delbrueckii subsp. lactis DSM 7290, and partial characterization of the enzyme

et al. 1994

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.Universitat Kaiserslautern, Fac h be reic h B iolog ie, Abteilung Mikrobiologie, Postfach 3049, 67653 Kaisersla ut er n, GermanyCell extracts of Lactobacillus delbrueckii subsp. lactis DSM 7290 were found to exhibit unique peptolytic ability against unusual fi-alanyl-dipeptides. In order to clone the gene encoding this activity, designated pepV, a gene library of strain DSM 7290 genomic DNA, prepared in the low-copy-number plasmid pLG339, was screened for heterologous expression in Escherichia coli. Recombinant clones harbouring pepV were identified by their ability to allow the utilization of carnosine (B-alanyl-histidine) as a source of histidine by the E. coli mutant strain UK197 (pepD, hisG). Complementation was observed in a colony harbouring a recombinant plasmid (pKVIOI), carrying pepV. A 2.4 kb fragment containing pepV was subcloned and its nucleotide sequence revealed an open reading frame (ORF) of 1413 nucleotides, corresponding to a protein with predicted molecular mass of 51 998 Da. A single transcription initiation site 71 bp upstream of the ATG translational start codon was identified by primer extension. No significant homology was detected between pepV or its deduced amino acid sequence with any entry in the databases. The only similarity was found in a region conserved in the ArgE/DapE/CPG2/YscS family of proteins. This observation, and protease inhibitor studies, indicated that pepV is of the metalloprotease type. A second ORF present in the sequenced fragment showed extensive homology to a variety of amino acid permeases from E. coli and Saccharomyces cerevisiae.

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“…Much information has accumulated on the role of proteolytic (caseinolytic) enzymes in the growth and nutrition of group N streptococci since this is of concern to the dairy industry (Thomas & Pritchard, 1987). However, knowledge concerning casein dissimilation is, for a number of reasons, relevant to other organisms including the oral streptococci.…”

Section: Introductionmentioning

confidence: 99%

The utilization of casein and amino acids by Streptococcus sanguis P4A7 in continuous culture

Rogers

Reynolds²

1990

Journal of General Microbiology

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Streptococcus sang& P4A7 was grown in glucose limited conditions in continuous culture at pH 7-0 in a chemically defined medium containing either free amino acids or casein as the organic nitrogen source. Apart from aspartate and threonine, which were poorly utilized at the higher dilution rates, all amino acids in the free-amino-acid medium were utilized to various extents. At the higher dilution rates, aspartate actually increased in concentration, probably due to deamidation of asparagine. The amino acid most utilized at all dilution rates was arginine, with up to 99 yo of the amino acid being consumed. Both casein and its aS1 -casein fraction supported growth at a level only slightly lower than that obtained with the free-amino-acid medium, provided that either cysteine or thioglycollate was present. With the exceprion of tyrosine, nearly all of the amino acyl residues ofaSl -casein were utilized to some degree. In general, the higher the concentration of each amino acid in the medium (whether free or as part of as1-casein) the higher the extent of utilization by S. sanguis P4A7. Only 50% of the arginyl residues (0-16 mM) of aslcasein were utilized compared with 99% of free arginine (1.5 mM) under similar conditions, suggesting that only 50% of the aS1 -casein arginine was accessible to the organism. From a comparison of the amino acid composition of a,,-casein with that of the high M, fraction of the culture supernatant it was concluded that leucine, phenylalanine, lysine, histidine, arginine and valine were specifically released from a,, -casein by the endo-and exopeptidase activity of S. sanguis P4A,.

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Proteolytic enzymes of dairy starter cultures

Cited by 306 publications

References 88 publications

Purification and characterization of the free form of the lactococcal extracellular proteinase and its autoproteolytic cleavage products

Purification and characterization of the free form of the lactococcal extracellular proteinase and its autoproteolytic cleavage products

Cloning and nucleotide sequence analysis of pepV, a carnosinase gene from Lactobacillus delbrueckii subsp. lactis DSM 7290, and partial characterization of the enzyme

The utilization of casein and amino acids by Streptococcus sanguis P4A7 in continuous culture

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