Hydrolysis of 14 C-labelled proteins by rumen micro-organisms and by proteolytic enzymes prepared from rumen bacteria

Abstract

1. Proteins were labelled with 14 C in a limited reductive methylation using [ 14 C]formaldehyde and sodium borohydride. 2. The rate of hydrolysis of purified proteins was little (< 10%) affected by methylation and the 14 C-labelled digestion products were not incorporated into microbial protein during a 5 h incubation with rumen fluid in vitro. It was therefore concluded that proteins labelled with 14 C in this way are valid substrates for study with rumen micro-organisms. 3. The patterns of digestion of 14 C-labelled fish meal, linseed meal and groundnut-protein meal by rumen micro-organisms in vitro were similar to those found in vivo. 4. The rates of hydrolysis of a number of 14 C-labelled proteins, including glycoprotein II and lectin from kidney beans ( Phaseolus vulgaris ), were determined with mixed rumen micro-organisms and with proteases extracted from rumen bacteria. Different soluble proteins were digested at quite different rates, with casein being most readily hydrolysed. 5. Proteins modified by performic acid oxidation, by cross-linking using 1,6-di-iso-cyanatohexane or by diazotization were labelled with 14 C. Performic acid treatment generally increased the susceptibility of proteins to digestion, so that the rates of hydrolysis of performic acid-treated proteins were more comparable than those of the unmodified proteins. Cross-linking resulted in a decreased rate of hydrolysis except with the insoluble proteins, hide powder azure and elastin congo red. Diazotization had little effect on the rate of hydrolysis of lactoglobulin and albumin, but inhibited casein hydrolysis and stimulated the breakdown of γ-globulin.