PMCC PMCC

Search tips
Search criteria

Advanced
Results 1-6 (6)
 

Clipboard (0)
None

Select a Filter Below

Journals
Authors
more »
Year of Publication
Document Types
1.  Involvement of enzyme-substrate charge interactions in the caseinolytic specificity of lactococcal cell envelope-associated proteinases. 
Applied and Environmental Microbiology  1995;61(11):3934-3939.
Three series of oligopeptides were synthesized to investigate the proposal that a major factor in determining the differences in specificity of the lactococcal cell surface-associated proteinases against caseins is the interactions between charged amino acids in the substrate and in the enzyme. The sequences of the oligopeptides were based on two regions of kappa-casein (residues 98 to 111 and 153 to 169) which show markedly different susceptibilities to PI- and PIII-type lactococcal proteinases. In each series, one oligopeptide had an identical sequence to that of the kappa-casein region, while in the others, one or more charged residues were substituted by an amino acid of opposite charge, i.e., His<-->Glu. Generally, substitution of His by Glu in the oligopeptides corresponding to residues 98 to 111 of kappa-casein resulted in reduced cleavage of susceptible bonds by the PI-type proteinase and increased cleavage of susceptible bonds by the PIII-type proteinase. In the case of the oligopeptide corresponding to residues 153 to 169 of kappa-casein, one major cleavage site was evident, and the bond was hydrolyzed by both types of proteinase (even though this sequence in kappa-casein itself is extremely resistant to the PI-type enzyme). Substitution of Glu by His in this oligopeptide, even in the P7 position, resulted in increased cleavage of the bond by the PI-type proteinase and reduced cleavage by the PIII-type proteinase. C-terminal truncation of this oligopeptide resulted in a 100-fold decrease in the rate of hydrolysis of the susceptible bond and a change in the pattern of cleavage.(ABSTRACT TRUNCATED AT 250 WORDS)
PMCID: PMC167699  PMID: 8526506
2.  Occurrence of Arginine Deiminase Pathway Enzymes in Arginine Catabolism by Wine Lactic Acid Bacteria 
l-Arginine, an amino acid found in significant quantities in grape juice and wine, is known to be catabolized by some wine lactic acid bacteria. The correlation between the occurrence of arginine deiminase pathway enzymes and the ability to catabolize arginine was examined in this study. The activities of the three arginine deiminase pathway enzymes, arginine deiminase, ornithine transcarbamylase, and carbamate kinase, were measured in cell extracts of 35 strains of wine lactic acid bacteria. These enzymes were present in all heterofermentative lactobacilli and most leuconostocs but were absent in all the homofermentative lactobacilli and pediococci examined. There was a good correlation among arginine degradation, formation of ammonia and citrulline, and the occurrence of arginine deiminase pathway enzymes. Urea was not detected during arginine degradation, suggesting that the catabolism of arginine did not proceed via the arginase-catalyzed reaction, as has been suggested in some earlier studies. Detection of ammonia with Nessler's reagent was shown to be a simple, rapid test to assess the ability of wine lactic acid bacteria to degrade arginine, although in media containing relatively high concentrations (>0.5%) of fructose, ammonia formation is inhibited.
PMCID: PMC1388333  PMID: 16534912
3.  Fructose 1,6-diphosphate-activated L-lactate dehydrogenase from Streptococcus lactis: kinetic properties and factors affecting activation. 
Journal of Bacteriology  1977;131(1):82-91.
The L-(+)-lactate dehydrogenase (L-lactate:NAD+ oxidoreductase, EC 1.1.1.27) of Streptococcus lactis C10, like that of other streptococci, was activated by fructose 1,6-diphosphate (FDP). The enzyme showed some activity in the absence of FDP, with a pH optimum of 8.2; FDP decreased the Km for both pyruvate and reduced nicotinamide adenine dinucleotide (NADH) and shifted the pH optimum to 6.9. Enzyme activity showed a hyperbolic response to both NADH and pyruvate in all the buffers tried except phosphate buffer, in which the response to increasing NADH was sigmoidal. The FDP concentration required for half-maximal velocity (FDP0.5V) was markedly influenced by the nature of the assay buffer used. Thus the FDP0.5V was 0.002 mM in 90 mM triethanolamine buffer, 0.2 mM in 90 mM tris(hydroxymethyl)aminomethanemaleate buffer, and 4.4 mM in 90 mM phosphate buffer. Phosphate inhibition of FDP binding is not a general property of streptococcal lactate dehydrogenase, since the FDP0.5V value for S. faecalis 8043 lactate dehydrogenase was not increased by phosphate. The S. faecalis and S. lactis lactate dehydrogenases also differed in that Mn2+ enhanced FDP binding in S. faecalis but had no effect on the S. lactis dehydrogenase. The FDP concentration (12 to 15 mM) found in S. lactis cells during logarithmic growth on a high-carbohydrate (3% lactose) medium would be adequate to give almost complete activation of the lactate dehydrogenase even if the high FDP0.5V value found in 90 mM phosphate were similar to the FDP requirement in vivo.
PMCID: PMC235394  PMID: 17595
4.  Regulation of the L-lactase dehydrogenase from Lactobacillus casei by fructose-1,6-diphosphate and metal ions. 
Journal of Bacteriology  1975;121(3):777-784.
The lactate dehydrogenase of Lactobacillus casei, like that of streptococci, requires fructose-1,6-diphosphate (FDP) for activity. The L. casei enzyme has a much more acidic pH optimum (pH 5.5) than the streptococcal lactate dehydrogenases. This is apparently due to a marked decrease in the affinity of the enzyme for the activator with increasing pH above 5.5; the concentration of FDP required for half-maximal velocity increase nearly 1,000-fold from 0.002 mM at pH 5.5 to 1.65 mM at 6.6. Manganous ions increase the pH range of activity particularly on the alkaline side of the optimum by increasing the affinity for FDP. This pH dependent metal ion activation is not specific for Mn2+. Other divalent metals, Co2+, Cu2+, Cd2+, Ni2+, Fe2+, Fe2+, and Zn2+ but not Mg2+, will effectively substitute for Mn2+, but the pH dependence of the activation differs with the metal ion used. The enzyme is inhibited by a number of commonly used buffering ions, particularly phosphate, citrate, and tris (hydroxymethyl) aminomethane-maleate buffers, even at low buffer concentrations (0.02 M). These buffers inhibit by affecting the binding of FDP.
PMCID: PMC246003  PMID: 234946
5.  Production of Staphylococcal Enterotoxins A, B, and C Under Conditions of Controlled pH and Aeration 
Infection and Immunity  1973;7(6):847-854.
The production of enterotoxins A, B, and C by nine strains of Staphylococcus aureus has been studied under controlled conditions in a fermenter. The strain to strain differences between staphylococci producing a specific enterotoxin were very marked. Increasing aeration in shake flasks improved both growth and production of all extracellular proteins measured other than that of enterotoxin C, the yield of which was decreased in one strain at high aeration. Silicone antifoam decreased the production of extracellular proteins, although enterotoxin A production from three strains was much less affected than that of enterotoxins B and C. In a detailed study of three strains, production of enterotoxins A and C was considerably greater in a defined amino acid medium than in a casein hydrolysate medium and was optimal for all three enterotoxins between pH 6.5 and 7.3. Changes in the pH or medium used in the fermenter that led to increased enterotoxin production could generally be correlated with a change in growth pattern, showing an extended transition period from exponential to stationary phase. Three out of five enterotoxin-A producing strains produced significantly more enterotoxin at a controlled pH of 6.5 in the fermenter than in shake-flask cultures. The yields with strain 100 were about five times greater than hitherto reported. Since many foods are buffered at pH 6 to 6.5, some strains may, therefore, produce sufficient enterotoxin A to cause food poisoning, although little or none may be produced when grown under normal testing procedures.
PMCID: PMC422773  PMID: 4716543
6.  Specificity of hydrolysis of bovine kappa-casein by cell envelope-associated proteinases from Lactococcus lactis strains. 
The cell envelope-associated proteinases from Lactococcus lactis subsp. cremoris H2 (a PI-type proteinase-producing strain) and SK11 (a PIII-type proteinase-producing strain) both actively hydrolyze the kappa-casein component of bovine milk but with significant differences in the specificity of peptide bond hydrolysis. The peptide bonds Ala-23-Lys-24, Leu-32-Ser-33, Ala-71-Gln-72, Leu-79-Ser-80, Met-95-Ala-96, and Met-106-Ala-107 were cleaved by both proteinase types, although the relative rates of hydrolysis at some of these sites were quite different for the two proteinases. Small histidine-rich peptides were formed as early products of the action of the cell envelope-associated proteinases on kappa-casein, implicating this casein as a possible significant source of histidine, which is essential for starter growth. The major difference between the two proteinase types in their action on kappa-casein was in their ability to cleave bonds near the C-terminal end of the molecule. The bond Asn-160-Thr-161 and, to a lesser extent, the bond Glu-151-Val-152 were very rapidly cleaved by the PIII-type proteinase, whereas hydrolysis of these bonds by the PI-type proteinase was barely detectable (even after 24 h of digestion). Differential hydrolysis of kappa-casein at these sites by the two different proteinase types resulted in the formation of distinctive, high-M(r) products detectable by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.(ABSTRACT TRUNCATED AT 250 WORDS)
Images
PMCID: PMC201395  PMID: 8161175

Results 1-6 (6)