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Biomedical subjects

J Kay

Publications and source records attributed to J Kay.

At least 271 records · Page 15Linked to original sources
PubMed1979

A possible role for neutral proteolysis in the degradation of intracellular proteins.

A neutral proteinase has been solubilized from rat intestinal muscle by extraction at low ionic strength. It has an apparent mol. wt. of 33,000 and is stable only around neutral pH. Characterization studies with specific inhibitors and substrates have shown it to be a trypsin-like serine proteinase. The rat proteinase and bovine pancreatic trypsin have equivalent activities as measured with peptide and denatured protein substrates but the rat proteinase is about 300 times more active than an equimolar amount of trypsin towards proteins in their native conformation. It has been shown that the activity of the rat proteinase can be modulated (1) by changing the conformation of the substrate protein(s) and (2) by means of an endogenous inhibitor. The inhibitor has been purified to homogeneity from rat intestinal muscle. It has a mol. wt. of 8,000 and binds only weakly to the rat proteinase (Ki approximately equal to 10(-6) M). It did not inhibit any of the other proteinases tested. The implications for such a proteinase--inhibitor system in the non-lysosomal pathway of intracellular protein degradation are considered.

Animals↗
PubMed1978

The susceptibility of muscle phosphorylases a and b to digestion by a neutral proteinase from rat intestinal muscle. Comparison with the effects produced by pancreatic trypsin and chymotrypsin.

1. Phosphorylase b was inactivated three times more rapidly than phosphorylase a by a neutral, trypsin-like proteinase from rat intestinal muscle. Digestion of phosphorylase a produced a modified form which was deactivated by AMP. Removal of the pyridoxal phosphate cofactor increased the rate of inactivation of the b form by about 3-fold but the subceptibility of apophosphorylase a was no different from the holo form. 2. The extent of proteolysis of both holoenzyme forms, as guaged by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, was limited and similar digestion patterns were obtained in both cases. 3. With (32)P-labelled phosphorylase a as substrate, the initial event in the inactivation was the release of a trichloroacetic acid-soluble peptide from the N-terminus of the enzyme, leaving the original 100000 subunit form essentially unchanged. Subsequent proteolysis was restricted, producing derivatives of mol.wt. 85000, 70000 and 65000, none of which contained any radioactive label. 4. By treatment of inactivated phosphorylase b with carboxypeptidase B, it was shown that the intestinal muscle proteinase had cleaved approximately 3 -Lys-X and 3 -Arg-X bonds in the polypeptide. 5. The protective effects of various allosteric modulators of phosphorylase on the inactivation of the a and b forms were generally in agreement with the known roles of the modifiers. Glucose increased the susceptibility of phosphorylase a. 6. Inactivation of phosphorylase b by trypsin and chymotrypsin also resulted in limited proteolysis but, in both cases, the digestion patterns obtained on sodium dodecyl sulphate/polyacrylamide gels were different from each other and from the pattern obtained with the intestinal muscle proteinase. 7. Inactivation of phosphorylase b by the muscle proteinase is about 100 times more rapid than the effects produced by trypsin or chymotrypsin when the activities are compared on an equimolar basis. 8. Consideration is given to regulation of the rate of enzyme degradation intracellularly by modulation of the conformation and susceptibility of the enzyme via factors such as covalent modification, allosteric ligands and state of aggregation.

Adenosine Monophosphate↗
PubMed1978

The inactivation of native enzymes by a neutral proteinase from rat intestinal muscle.

1. The solubilization and partial purification of a proteinase from the intestinal smooth muscle of rats fed on protein-free diets are described. 2. It has a mol.wt. of about 33000 and it is stable over a narrow pH range. 3. From its susceptibility to known modifers of proteolytic enzymes, it appears to be a serine proteinase of a trypsin-like nature. Active-site titration with soya-bean trypsin inhibitor shows that the concentration of proteinase was about 3 microgram/g wet wt. of intestinal smooth muscle. However, the muscle proteinase demonstrates a marked ability for inactivating enzymes in their native conformation at neutral pH. It is about 100 times more efficient than pancreatic trypsin when the inactivating activities are compared on an approximately equimolar basis. 4. Inactivation of the substrate enzymes is accompanied by limited proteolysis, as demonstrated by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. 5. An endogenous inhibitor was separated from the proteinase by fractionation with (NH4)2SO4. 6. Contamination of the muscle tissue by lumen, mucosal or blood proteinases and inhibitors is shown to be unlikely. 7. A role for the neutral trypsin-like proteinase in initiating the degradation of intracellular enzymes is considered.

Animals↗
PubMed1978

Terminating the doctor-patient relationship.

This paper addresses a neglected area of medical student education--terminating the doctor-patient relationship. Rich clinical teaching opportunities are lost when educators fail to illustrate the significant emotional aspects of ending this highly personal affect-laden experience. The author explores these teaching opportunities through examination of elements of doctor-patient relationships, professional development, unresolved doctor-patient conflicts, role underevaluation, patient gifts, and referral procedures.

Conflict, Psychological↗
PubMed1977

The first step in the activation of chicken pepsinogen is similar to that of prochymosin.

Chicken pepsinogen was incubated at pH2.5 with pepstatin. The zymogen activated itself by a sequential mechanism and an intact peptide derived from residues 1-26 in the protein was released in the first step. This peptide was found to inhibit the milk-clotting activities of pig and chicken pepsins and calf chymosin but to different extents.

Amino Acid Sequence↗
PubMed1977

The first cleavage site in pepsinogen activation.

By incubation of porcine, bovine, canine, or chicken pepsinogens and calf prochymosin with pepstatin at pH 2.5, the first active protein generated on activation is trapped in an inactive complex. The first activation peptide liberated from porcine pepsinogen has been identified as residues 1-16 whereas that from prochymosin is derived from residues 1-27. This suggests that pepsin and chymosin are not formed by one-step conversions from their zymogens, but by (different) sequential, activation mechanisms.

Amino Acid Sequence↗
PubMed1977

A neutral protease from rat intestinal muscle. A possible role in the degradation of native enzymes.

A membrane-limited protease has been solubilised and partially purified from the intestinal smooth muscle of rats fed on protein free diets. This neutral protease has a mol. wt. of around 33,000 and from its susceptibility to several known modifiers of proteolytic enzymes, it appears to be trypsin-like. It is stable over a relatively narrow pH range and it appears to have a markedly enhanced ability over trypsin for inactivating substrate enzymes in their native conformations through limited proteolysis. The rate of inactivation of substrate enzymes can be modulated by cofactors, allosteric ligands, or by changes in ionic strength. In addition, a specific protein inhibitor of the protease has been measured and levels of this are high in animals fed on normal diets. On administration of protein free diets, the inhibitory activity is depleted. Contamination of the muscle tissue by lumenal, mucosal or blood proteases and inhibitors has been excluded. A role for the neutral protease in initiating the turnover of intracellular enzymes is postulated.

Animals↗
PubMed1977

The susceptibility of glycogen phosphorylase to inactivation by endogenous and exogenous proteases.

Phosphorylases a and b were inactivated very rapidly by a neutral, trypsin-like protease from rat intestinal muscle. With 32P-phosphorylase a as substrate, it was shown that the initial event in the inactivation was the release of a small, phosphopeptide from the N-terminus of the enzyme, leaving the original 100,000 subunit form virtually unchanged. Subsequent proteolysis was very limited, producing 85, 70 and 65,000 mol. wt. derivatives. The effects of several allosteric modulators of phosphorylase on the rates of inactivation of the two enzymes were studied. Removal of the pyridoxal phosphate cofactor from phosphorylase increased the susceptibility of the b form by three fold while the a form was unaffected. By comparison of these effects with those obtained from digestion with trypsin and chymotrypsin, it is concluded that the intestinal muscle protease has a markedly enhanced ability for inactivating enzymes in their native conformation. Assuming that this property is reflected in vivo, a possible role such neutral proteases in initiating protein degradation is advanced.

Animals↗