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Homology modelling of cell-envelope proteinase

Homology modelling of cell-envelope proteinase. Engineering of the substrate binding region of the subtilisin-like, cell-envelope proteinase of Lactococcus lactis Protein Engineering 6 (1993) 927-937.

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Homology modelling of cell-envelope proteinase

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  1. Homology modelling of cell-envelope proteinase Engineering of the substrate binding region of the subtilisin-like, cell-envelope proteinase of Lactococcus lactis Protein Engineering 6 (1993) 927-937 Roland Siezen, Paul Bruinenberg, Pieter Vos, Ingrid van Alen-Boerrigter, Monique Nijhuis, Arno Alting, Fred Exterkate, and Willem de Vos

  2. Flavour formation in cheese Milk Lactose/ citric acid Casein Fat chymosin protease lipases esterases metabolic enzymes peptides peptidases fatty acids pyruvate amino acids metabolic enzymes amino acid convertases flavour compounds

  3. Proteolysis by lactic acid bacteria Casein peptides PepA PepX PrtP PepN L.lactis

  4. +8 +151 I-domain +14 +15 +28 PrtP protease domain C 166 N Ser Asp His Ca1 substrate binding cleft 137-139 Ca2

  5. as1-casein(1-23) degradation by PrtPR-P-K-H-P-I-K-H-Q-G-L-P-Q-E-V-L-N-E-N-L-L-R-F 5 10 15 20 wild-type N166D AKT(137-139)GDT AKT(137-139)GPP AKT(137-139)GLA D(238-388) Main cleavage sites: pH 6.5, low NaCl

  6. Conclusions Homology modelling • substrate-binding region of PrtP can be modelled • several (large) loops cannot be modeled • model suffices for general predictions Protein engineering • mutations in substrate-binding region, based on homology modelling • mutants show altered proteolytic specificity • tested mutants under cheese making conditions

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