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C Abell

Publications and source records attributed to C Abell.

48 records · Page 3Linked to original sources
PubMed1990

Investigation of putative active-site lysine residues in hydroxymethylbilane synthase. Preparation and characterization of mutants in which (a) Lys-55, (b) Lys-59 and (c) both Lys-55 and Lys-59 have been replaced by glutamine.

A new construct carrying the hemC gene was transformed into Escherichia coli, resulting in approx. 1000-fold over-expression of hydroxymethylbilane synthase (HMBS). This construct was used to generate HMBS in which (a) Lys-55, (b) Lys-59 and (c) both Lys-55 and Lys-59 were replaced by glutamine (K55Q, K59Q and K55Q-K59Q respectively). All three modified enzymes are chromatographically separable from wild-type enzyme. Kinetic studies showed that the substitution K55Q has little effect whereas K59Q causes a 25-fold decrease in Kapp. cat./Kapp. m. Treatment of K55Q, K59Q and K55Q-K59Q separately with pyridoxal 5'-phosphate and NaBH4 resulted in incomplete and non-specific reaction with the remaining lysine residues. Pyridoxal modification of Lys-59 in the K55Q mutant caused greater enzymic inactivation than similar modification of Lys-55 in K59Q. The results in sum show that, though Lys-55 and Lys-59 may be at or near the active site, neither is indispensable for the catalytic activity of HMBS.

Binding Sites↗
PubMed1989

Isolation and characterisation of a cDNA clone for a chlorophyll synthesis enzyme from Euglena gracilis. The chloroplast enzyme hydroxymethylbilane synthase (porphobilinogen deaminase) is synthesised with a very long transit peptide in Euglena.

A cDNA expression library was constructed from light-grown Euglena gracilis poly(A)-rich RNA in lambda gt11. Antibodies to Euglena hydroxymethylbilane synthase, the third enzyme in the porphyrin biosynthetic pathway, were used to screen the library and a clone encoding part of the sequence of hydroxymethylbilane synthase was identified. This was used to rescreen the library and a full-length clone was isolated, which encoded not only the entire mature protein (Mr 36,927), but also an N-terminal extension of 139 amino acids. The deduced Mr of the whole polypeptide is 51,744, which corresponds to the size of the protein immunoprecipitated from the translation products of Euglena poly(A)-rich RNA. The mature protein is 60-70% similar to hydroxymethylbilane synthase from human erythrocytes and Escherichia coli. The sequence of the N-terminal extension has similarities to both the transit peptides of chloroplast proteins and those for the endoplasmic reticulum. This is the first report both of a cDNA clone for an enzyme of the chlorophyll biosynthetic pathway and of a putative transit peptide for a nuclear-encoded Euglena protein.

Amino Acid Sequence↗
PubMed1989

Evidence that pyridoxal phosphate modification of lysine residues (Lys-55 and Lys-59) causes inactivation of hydroxymethylbilane synthase (porphobilinogen deaminase).

A recombinant strain of Escherichia coli has been constructed that produces approx. 200 times the amount of hydroxymethylbilane synthase found in wild-type E. coli [Hart, Abell & Battersby (1986) Biochem. J. 240, 273-276]. Enzyme purified from this strain is shown to be permanently inactivated by pyridoxal 5'-phosphate/NaB1H3(3)H1. The inactivation is not complete despite the fact that approx. 1 mol of lysine residues is modified per mol of enzyme. Evidence is gained showing that (a) modification of one of two conserved lysine residues (Lys-55 or Lys-59) results in inactivation of hydroxymethylbilane synthase and (b) these lysine residues are present in or close to the active site.

Amino Acid Sequence↗
PubMed1986

Purification, N-terminal amino acid sequence and properties of hydroxymethylbilane synthase (porphobilinogen deaminase) from Escherichia coli.

Hydroxymethylbilane synthase (porphobilinogen deaminase) was purified to apparent homogeneity from Escherichia coli. The enzyme is a monomer of Mr approx. 40,000. The Km for porphobilinogen and relative Vmax. values have been obtained at various pH values over the range 6.2-8.8, enabling pK values for ionizable groups important for activity to be determined. The N-terminal amino acid sequence is presented.

Amino Acid Sequence↗
PubMedDate not supplied

Nonlinear analysis of partial dopamine agonist effects on cAMP in C6 glioma cells.

Most drugs have some efficacy so that improved methods to determine the relative intrinsic efficacy of partial agonists should be of benefit to preclinical and clinical investigators. We examined the effects of partial D(1) or partial D(2) dopamine agonists using a partial agonist interaction model. The dependent variable was the modulation of the dopamine-receptor-mediated cAMP response in C6 glioma cells selectively and stably expressing either D(1) or D(2) recombinant dopamine receptors. The dissociation constant (K(B)) and relative intrinsic efficacy (E(r)) for each partial agonist were calculated using a partial agonist interaction null model in which the effects of fixed concentrations of each partial agonist on the dopamine dose-response curve were evaluated. This model is an extension of the competitive antagonist null model to drugs with efficacy and assumes only that the log-dose--response curve is monotonic. Generally, the partial agonist interaction model fit the data, as well as fits of the independent logistic curves. Furthermore, the partial agonist K(B) values could be shared across partial agonist concentrations without worsening the model fit (by increasing the residual variance). K(B) values were also similar to drug affinities reported in the literature. The model was validated in three ways. First, we assumed a common tissue stimulus parameter (beta) and calculated the E(r) values. This provided a qualitative check on the interaction model results. Second, we calculated new relative efficacy values, E(r)(beta), using the beta estimate. Third, we calculated relative efficacy using relative maxima times midpoint shift ratios (J. Theor. Biol. 198 (1999) 347.). All three methods indicated that the present model yielded reasonable estimates of affinity and relative efficacy for the set of compounds studied. Our results provide a quick and convenient method of quantification of partial agonist efficacy. Special applications and limitations of the model are discussed. In addition, the present results are the first report of the relative intrinsic efficacy values for this set of D(2) ligands.

2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-ben↗
PubMedDate not supplied

Novel chemical tagging method for combinatorial synthesis utilizing suzuki chemistry and fourier transform ion cyclotron resonance mass spectrometry.

We describe a novel chemical tagging strategy for combinatorial solid-phase chemistry. The tags used are para-substituted alkyl phenols, with the first tags attached directly to the chloromethyl polystyrene and subsequent tags attached via Suzuki couplings using either aryl diboronic acids or aryl iodides. The identities of the tags attached to a single bead are discovered by the high-resolution, accurate mass technique of Fourier transform ion cyclotron resonance mass spectrometry. The method is exemplified for the coded assembly of a tripeptide.

Alkynes↗
PubMedDate not supplied

Teaching "family": evaluating a multidimensional clerkship curriculum.

BACKGROUND: A multidimensional program that includes workshops on family systems, family interviewing, a home visit, and faculty precepting at East Tennessee State University (ETSU) is described and evaluated. METHODS: Over a 3-year period, ETSU junior clerks received systematic instruction on family issues in medical practice. Evaluations were obtained at the conclusion of the experience. RESULTS: The number of students who rated the importance of family in family practice as "very strong" increased from 14 (28%) to 33 (66%) between the beginning and the end of the rotation. The majority of students reported that each of the curricular elements on family had a "strong positive impact" and rated home visits as most important in promoting their understanding of families. CONCLUSIONS: The curriculum on family had a significant role in enhancing students' appreciation of the importance of the family in medicine.

Clinical Clerkship↗