Purification and characterization of a pregastric esterase from a hygienized kid rennet paste.
Rennet pastes obtained by maceration of gastric tissues from suckling kids are used traditionally to produce some artisanal cheeses in Spain. Besides milk-clotting function, rennet pastes provide proteolytic activity and lipolytic system, essentially pregastric, necessary in the development of piquant flavor typical of these cheeses. A simple and reproducible procedure allows us to obtain a standardized rennet paste that posses the desired activity and is of good microbiological quality. Concomitantly, a kid pregastric esterase (KPGE) was purified to homogeneity. The purification procedure was based on an aqueous extract of hygienized rennet paste (HRP), which was chromatographed on DEAE-Sepharose Fast Flow then adsorbed on phenyl superose followed by a re-chromatography on the same column. The final enzymatic preparation, where the overall activity recovery was 3%, showed a molecular mass of 53 kDa. The highest activity was determined on p-nitrophenyl butyrate, but marked hydrolysis was also detected on beta-naphthyl caprylate. In contrast, low activity on tributyrin (substrate under emulsion form) was detected, thus confirming the esterase character of purified enzyme.
Calvo
M V
MV
Dpto. de Ciencia y Tecnología de Productos Lácteos, Instituto del Frío (CSIC), C/ José Antonio Novais, Ciudad Universitaria, Madrid, Spain. mcalvo@if.csic.es
Fontecha
J
J
D000818:Animals; D002611:Cheese; D012085:Chymosin / Q000737:chemistry*; D005511:Food Handling; D006041:Goats; D006863:Hydrogen-Ion Concentration; D008049:Lipase / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D013270:Stomach / Q000201:enzymology; D013379:Substrate Specificity; D013696:Temperature
C025350:rennet; D008049:Lipase; C043101:gastric triacylglycerol lipase; D012085:Chymosin