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Biomedical subjects

M Knight

Publications and source records attributed to M Knight.

At least 109 records · Page 6Linked to original sources
PubMed1972

A heat-stable nicotinamide-adenine dinucleotide glycohydrolase from Pseudomonas putida KB1. Partial purification and some properties of the enzyme and an inhibitory protein.

A thermostable NAD(P)(+) glycohydrolase (EC 3.2.2.6) detected in cell-free extracts of Pseudomonas putida KB1 was purified to a single component on polyacrylamide-gel electrophoresis. A heat-labile inhibitor of the enzyme was also partially purified. Enzyme free of inhibitor is present in culture supernatants. After an ultrasonic treatment enzyme-inhibitor complex and excess of inhibitor are present in both the cell-debris and soluble fractions. The general properties of the enzyme and inhibitor are described. The molecular weights of enzyme, inhibitor and enzyme-inhibitor complex, determined by gel filtration are about 23500, 15000 and 35000 respectively. The binding of inhibitor and enzyme is inhibited by the presence of substrate.

Cell-Free System↗
PubMed1966

The synthesis of amino acids by Methanobacterium omelianskii.

1. Methanobacterium omelianskii was grown on (14)CO(2) and unlabelled ethanol, or on [1-(14)C]- or [2-(14)C]-ethanol and unlabelled carbon dioxide. The cell protein was hydrolysed and certain of the amino acids were isolated and degraded. 2. Carbon from both carbon dioxide and ethanol is used for biosynthesis of amino acids, and in most cases ethanol is incorporated as a C(2) unit. Ethanol carbon atoms and carbon dioxide carbon atoms apparently enter the same range of compounds. Ethanol and carbon dioxide are equally important as sources of cell carbon. 3. The origins of carbon atoms of aspartate, alanine, glycine, serine and threonine are consistent with the synthesis of these amino acids, by pathways known to exist in aerobic organisms, from pyruvate arising by a C(2)+C(1) condensation. The proportion of total radioactivity found in C-1 of lysine, proline, methionine and valine is consistent with synthesis of these amino acids by pathways similar to those found in Escherichia coli. Isoleucine is probably formed by carboxylation of a C(5) precursor formed entirely from ethanol. Glutamate is formed by an unknown pathway.

Amino Acids↗
PubMedDate not supplied

Brain endopeptidase generates enkephalin from striatal precursors.

An enzyme capable of converting putative opioid peptide intermediates to free enkephalin has been purified 300-fold from washed rat brain membranes. The action of this enzyme, an enkephalin-generating endopeptidase (EGE), was compared with the action of carboxypeptidase B after trypsin treatment on enkephalin precursor peptides present in rat striata. After Sephadex G-100 gel filtration of striatal material, fractions were radioimmunoassayed for enkephalin content using an antiserum specific for the carboxyl terminal of enkephalin. Additionally, aliquots of the column fractions were treated with either trypsin and carboxypeptidase B, trypsin and EGE, or EGE alone. The peak of enkephalin immunoreactivity increased with the enzymes' treatment indicating the conversion of the low molecular weight proenkephalin precursor peptides to enkephalin. Trypsin and EGE generated almost as much enkephalin as trypsin and carboxypeptidase B in the conditions of the experiment. Thus EGE is capable of processing precursors to enkephalin after the action of trypsin-like enzyme(s) in the brain. The gel filtration fractions containing enkephalin and its low molecular weight precursors were pooled and one-half treated with EGE. The contents were analyzed by HPLC and the increase in immunoreactivity co-eluted with enkephalin and Leu-enkephalin. Small peptides found to be the most potent competitive inhibitors of this enzyme are Met-Arg-Phe-Ala, and Met-Arg-Phe.

Amino Acid Sequence↗
PubMedDate not supplied

Tolerance to the anti-avoidance properties of cholecystokinin-octapeptide.

Previous investigations have suggested a neuroleptic-like action of cholecystokinin-octapeptide (CCK8) on conditioned-avoidance behavior. This study was initiated to test tolerance to this effect. Rats were trained to avoid electric shock in a shuttle box under a free-operant (Sidman) avoidance paradigm. Each shuttle response postponed a 0.2 sec, 1 mA shock for 20 sec. If the rat failed to respond, shock was delivered every 5 sec until a response occurred. After avoidance training, half of the rats received two daily injections of CCK8 (0.320 mg/kg, IP) and half received saline for 7 days. Rats were then tested on the Sidman avoidance 1 min after receiving CCK8 (0.640 mg/kg, IP) or saline. CCK8 depressed avoidance responding if rats received saline for 7 days prior to the test. Rats pretreated with CCK8 for 7 days were not significantly affected by CCK8 during the avoidance test. Thus, repeated injections of CCK8 result in tolerance to its anti-avoidance properties.

Animals↗
PubMedDate not supplied

Cholecystokinin octapeptide analogues stable to brain proteolysis.

Based on recent findings identifying the initial degradative cleavage of CCK-8 at the Met3-Gly4 bond by a metalloendopeptidase, two analogues of CCK-8 with D-Ala and D-Trp substitutions at the Gly4 position were synthesized as stable analogues. Their stability to proteolysis by brain membranes and their binding potency at central CCK receptors were quantified. Both peptides are stable to degradation by peptidases in cortical synaptic membrane preparations. The analogues are nearly equipotent to CCK-8 in their affinities for inhibition of 125I-CCK-33 binding to guinea pig cortical membranes. L-Ala and L-Trp substituted peptides were synthesized for comparison. Both these peptides are degraded by synaptic membranes and the L-Trp substituted peptide possesses a greatly reduced affinity for central CCK receptors. Therefore, the structure of CCK due to the D conformation of Gly is more capable of interacting with brain CCK receptors. Further conformational analysis will establish whether the stabilized structure is a beta-bend or a beta-turn. Since these peptides are highly potent and stable to brain proteolysis they may be useful as stable CCK analogues for in vivo application.

Animals↗