Bovine milk hydrophobic fraction of proteose-peptone was prepared by hydrophobic interaction fast protein liquid chromatography. This method has several advantages such as high rapidity, simple good reproducibility and less denaturation. The proteose-peptone was eluted from a TSK-Phenyl-5PW column with a 1 M-0 M ionic strength gradient of NaH 2 PO 4 , pH 6-8, using a 6 ml/min flow rate for 56 min. The quantity of protein injected was 62-5 mg; however, it could be increased up to 100 mg. The elution order was /J-CN-4P

No abstract

The influence of NaCl on bovine cY,,-casein (o&N) hydrolysis by papain (EC 3.4.22.2) was studied at pH 6.5 and 40°C. The electrophoretic analysis in presence of sodium dodecyl sulfate (SDS), showed that o&N was sequentially hydrolyzed as the reaction time increased. The analysis by isoelectric focusing electrophorcsis showed that after 1 min, the resulting peptides had their isoelectric point (PI) assessed between 4 and 9. With the increase of NaCl concentration, two peptides of apparent pI near pH 5 and 8.5 were progressively liberated. The chromatographic analysis by reverse phase HPLC showed that a major peak, observed after 1 min of reaction, persisted in the hydrolysates obtained in the presence of salt at all reaction times. It was concluded by the peptidic sequence determination that the hydrophobic C-terminal region of a,,-CN was not hydrolyzed in the presence of NaCI.

The influence of cu,,-casein concentration on the hydrolytic activity of papain was studied. High substrate concentration was inhibitory. In presence of NaCI, the papain affinity for cy,,-casein increased (the apparent Michaelis constant (K,. app.) decreased from 9.4 10e5 to 5.6 1O-5 M) and was lower for 0.08M NaCI. The maximum velocity (V,,, app.) was independant from NaCl concentration below 0.34M but decreased above. The enzyme-substrate affinity was increased by the addition of urea, but the V,, app. decreased. The inhibitory effect of excess substrate was more important with the presence of both urea and NaCl. Acetylation of a,,-casein showed that K,. app. (21.2 lo-jM) was independant of salt concentration, while the V,, app. varied, and the substrate inhibitory activity was suppressed.