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Biomedical subjects

R S Alexander

Publications and source records attributed to R S Alexander.

At least 37 records · Page 2Linked to original sources
PubMed1991

Engineering the hydrophobic pocket of carbonic anhydrase II.

Wild-type and mutant human carbonic anhydrases II, where mutations have been made in the hydrophobic pocket of the active site, have been studied by X-ray crystallographic methods. Specifically, mutations at Val-143 (the base of the pocket) lead to significant changes in catalytic activity and protein structure. The obliteration of a well-defined pocket in the Val-143----Phe and Val-143----Tyr mutants results in significantly diminished enzyme activity [(5 x 10(4))-fold and (3 x 10(5))-fold, respectively]; however, the activity of the Val-143----His mutant is diminished less (10(2)-fold), and deepening the pocket in the Val-143----Gly mutant results in only a 2-fold decrease in activity [Fierke et al., 1991 (preceding paper in this issue)]. These results indicate that the hydrophobic pocket is important for substrate association with the enzyme, but there are probably several catalytically acceptable substrate trajectories through this region of the enzyme structure. Additionally, each mutant protein exhibits long-range (ca. 10-15 A) compensatory structural changes which accommodate the Val-143 substitution. As such, the genetic-structural approach represented in this work serves as a three-dimensional paradigm for the redesign of specificity pockets in other protein catalysts.

Binding Sites↗
PubMed1991

Conformational mobility of His-64 in the Thr-200----Ser mutant of human carbonic anhydrase II.

The three-dimensional structure of the Thr-200----Ser (T200S) mutant of human carbonic anhydrase II (CAII) has been determined by X-ray crystallographic methods at 2.1-A resolution. This particular mutant of CAII exhibits CO2 hydrase activity that is comparable to that of the wild-type enzyme with a 2-fold stabilization of the E.HCO3- complex and esterase activity that is 4-fold greater than that of the wild-type enzyme. The structure of the mutant enzyme reveals no significant local changes accompanying the conservative T200S substitution, but an important nonlocal structural change is evident: the side chain of catalytic residue His-64 rotates away from the active site by 105 degrees about chi 1 and apparently displaces a water molecule. The displaced water molecule is present in the wild-type enzyme; however, the electron density into which this water is built is interpretable as an alternate conformation of His-64 with 10-20% occupancy. The rate constants for proton transfer from the zinc-water ligand to His-64 and from His-64 to bulk solvent are maintained in the T200S variant; therefore, if His-64 is conformationally mobile about chi 1 and/or chi 2 during catalysis, compensatory changes in solvent configuration must sustain efficient proton transfer.

Carbonic Anhydrases↗
PubMed1990

Hydrogen bond stereochemistry in protein structure and function.

Fifty high resolution protein structures from the Brookhaven Protein Data Bank have been analyzed for recurring motifs in hydrogen bond stereochemistry. Although an exhaustive analysis of hydrogen bond statistics has been presented by Baker & Hubbard, a detailed stereochemical analysis of classical donor (N-H, O-H, or S-H) and acceptor (N:, O:, or S:) structure within proteins is lacking. Here, we describe the preferential hydrogen bond stereochemistry for the side-chains of glutamate and aspartate (carboxylate), glutamine and asparagine (carboxamide), arginine (guanidinium), histidine (imidazole/imidazolium), tryptophan (indole), tyrosine (phenolic hydroxyl), lysine (ammonium), serine and threonine (alkyl hydroxyl), cysteine (thiol), methionine (thioether) and cystine (disulfide). Preferential hydrogen bond stereochemistry is governed by (1) the electronic configuration of acceptor atoms, (2) the steric accessibility of donor atoms and (3) the conformation of amino acid side-chains. Applications of hydrogen bond stereochemistry are useful in the interpretation of protein structure, function and stability. Additionally, this stereochemistry is a prerequisite to the interpretation of protein-other molecule recognition and biological catalysis.

Amino Acids↗
PubMed1983

An educational experience evaluated: the Christchurch Clinical School of Medicine.

This study explored the response of recent graduates to their clinical training at the University of Otago, Christchurch Clinical School of Medicine. Graduates rated their competence at dealing with a number of clinical problems and the adequacy of teaching received in relation to their anticipated requirements as medical practitioners. They also recorded information concerning their personal response to training--role identity, stress and anxiety felt during the course, and their commitment to ongoing education and research following training. Overall the course was regarded positively. In the light of anticipated requirements as medical practitioners some curricular gaps were perceived and criticisms of the course identified. Graduates were critical of their teachers' abilities in demonstrating interviewing and interpersonal skills. Over 70% of graduates felt that they had little or no role during much of their undergraduate clinical training. Many graduates felt that they had little opportunity to carry out research during training, nor the motivation to undertake research subsequently, but they were committed to continuing education.

Attitude of Health Personnel↗
PubMed1981

Microembolization induced oxygen utilization impairment in the canine gracilis muscle.

Impaired peripheral oxygen utilization coinciding with an elevated cardiac index, venous oxygen tension, and serum lactate with the loss of reactive hyperemic response have been observed in a large series of resuscitated trauma patients. We tested the hypothesis that these clinical findings were due to an alteration of the microcirculation caused by embolization of intravascular particulate matter. To test this hypothesis, we used the bilateral pump-perfused, isolated canine gracilis muscle preparation which we subjected to microembolization with 15 micrometers polystyrene spheres. Prior to microembolization, oxygen consumption was flow-limited up to 6 ml min-1 (r = .928) and at higher flows, oxygen consumption was independent of flow. Following microembolization, the relationship of oxygen consumption and blood flow remained correlated (r = .893), but there was less oxygen consumption at any given flow rate (P less than .05). Imidazole, 30 mg kg-1, IP administered to prevent platelet aggregation, resulted in the return of oxygen utilization to the preembolization value. PVO2 of the microembolized muscle was significantly higher than in the contralateral muscle, which was abolished after imidazole administration. These data suggest that microembolization leads to an oxygen utilization defect similar to that observed in the resuscitated trauma patient. Since this defect was reversed by imidazole administration, a humoral mechanism in the microcirculatory bed may act to restrict oxygen utilization following microembolization and trauma.

Animals↗
PubMed1979

Oxygen consumption in canine skeletal muscle following massive saline infusion.

To study the effect of interstitial edema on tissue oxygen transport, we infused buffered isotonic saline solution at a rate of 200 ml/kg/hr into mongrel dogs for two hours, measuring oxygen delivery and consumption in a skinned, innervated hindlimb. Hematocrit dropped gradually to 10.8 +/- 3.9%. Muscle water content increased 13% (P less than 0.05). However, oxygen consumption did not change significantly from control of 0.228 +/- 0.029 ml/min/100 gm. Femoral venous oxygen tension fell from control of 62.6 +/- 2.6 mm Hg to 31.4 +/- 2.3 mm Hg. Mean arterial flow increased to nearly twice the control level of 12.5 +/- 1.1 ml/min/100 gm and fell gradually, with the sustained 50% drop in vascular resistance largely explained by expected decreases in blood viscosity. We conclude that interstitial edema did not cause a significant defect in canine skeletal muscle oxygen utilization.

Animals↗
PubMed1976

Series elasticity of urinary bladder smooth muscle.

Loops of rat bladder were stretched between pins in vitro, supported by a clamp that could be suddenly shortened by activation of a solenoid to achieve a quick release of tension. The series elasticity measured in this fashion was found to follow an exponential course and to be modified by the rate of release, indicating a minor viscous component. Tissue length decreased and series elastic stiffness appeared to increase with muscle contraction, but no alteration in series elasticity was evident when the data were related to the tension existing in the tissue at the moment of quick release. Inactivation of the contractile system by removing calcium ion with ethylene glycol-bis-(beta-aminoethylether)-N,N'-tetraacetate (EGTA) similarly did not alter series elasticity when it was related to the tension existing in the tissue. Series elasticity during the stress relaxation following a stretch, and during the contracting and relaxing phases of rhythmic contractions, was also determined by tissue tension. The conclusion drawn is that contractile cross bridges do not contribute to the series elasticity measured in bladder tissue.

Animals↗