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Full-Text Articles in Biochemistry, Biophysics, and Structural Biology

Molecular Genetic Analysis Of Terminal Steps In Bacteriochlorophyll A Biosynthesis: Characterization Of A Rhodobacter Capsulatus Strain That Synthesizes Geranylgeranoil-Esterified Bacteriochlorophyll A, David Bollivar, Shaojie Wang, James P. Allen, Carl E. Bauer Oct 1994

Molecular Genetic Analysis Of Terminal Steps In Bacteriochlorophyll A Biosynthesis: Characterization Of A Rhodobacter Capsulatus Strain That Synthesizes Geranylgeranoil-Esterified Bacteriochlorophyll A, David Bollivar, Shaojie Wang, James P. Allen, Carl E. Bauer

David Bollivar

Site-directed mutational analysis of the Rhodobacter capsulatus photosynthesis gene cluster was undertaken in order to identify and characterize genetic loci involved in bacteriochlorophyll a biosynthesis. A mutant in orf304 was shown to accumulate the tetrapyrrole intermediate "bacteriochlorophyllide a" which is a tetrapyrrole that has a bacteriochlorophyll a ring structure without the presence of an esterifying alcohol. A mutant in orf391 is shown to synthesize acteriochlorophyll a that is esterified with geranylgeraniol rather than the normal phytol. This latter result provides the first genetic confirmation that esterification of bacteriochlorophyllide a initially involves the addition of a geranylgeraniol group followed by sequential ...


Effect Of Charged Residue Substitutions On The Thermodynamics Of Signal Peptide-Lipid Interactions For The Escherichia Coli Lamb Signal Sequence., Jeffrey D. Jones, Lila Gierasch Sep 1994

Effect Of Charged Residue Substitutions On The Thermodynamics Of Signal Peptide-Lipid Interactions For The Escherichia Coli Lamb Signal Sequence., Jeffrey D. Jones, Lila Gierasch

Lila Gierasch

We have used tryptophan fluorescence spectroscopy to characterize the binding affinities of an Escherichia coli LamB signal peptide family for lipid vesicles. These peptides harbor charged residue substitutions in the hydrophobic core region. Titrations of peptides with vesicles composed of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine and 1-palmitoyl-2-oleoyl-sn-3-phosphoglycerol (65:35 mol%), in conjunction with evaluation of peptide dissociation rates from these vesicles, were used to determine binding parameters quantitatively. We find that under low ionic strength conditions, point mutations introducing negatively charged aspartate residues substantially reduce peptide affinity relative to the wild-type peptide. However, the difference between wild-type and mutant peptide affinities was much lower ...


Heterologous Expression Of The Bchm Gene Product From Rhodobacter Capsulatus And Demonstration That It Encodes S-Adenosyl-L-Methionine: Mg-Protoporhyrin Ix Methyltransferase, David Bollivar, Ze-Yu Jiang, Carl E. Bauer, Samuel I. Beale Aug 1994

Heterologous Expression Of The Bchm Gene Product From Rhodobacter Capsulatus And Demonstration That It Encodes S-Adenosyl-L-Methionine: Mg-Protoporhyrin Ix Methyltransferase, David Bollivar, Ze-Yu Jiang, Carl E. Bauer, Samuel I. Beale

David Bollivar

The bacteriochlorophyll biosynthesis gene, bchM, from Rlodobacter capsulatus was previously believed to code for a polypeptide involved in formation of the cyclopentone ring of protochlorophyllide from Mg-protoporphyrin IX monomethyl ester. In this study, R. capsulatus bchM was expressed in Escherichia coli and the gene product was subsequently demonstrated by enzymatic analysis to catalyze methylation of Mg-protoporphyrin IX to form Mg-protoporphyrin IX monomethyl ester. Activity required the substrates Mg-protoporphyrin IX and S-adenosyl-L-methionine. 14C-labeled product was formed in incubations containing 14C-methyl-labeled S-adenosyl-L-methionine. On the basis of these and previous results, we also conclude that the bchH gene, which was previously reported to ...


Protein S-Thiolation In Hepatocytes Stimulated By T-Butyl Hydroperoxide, Menadione, And Neutrophils, Yuh-Cherng Chai, S. Hendrich, James Thomas Mar 1994

Protein S-Thiolation In Hepatocytes Stimulated By T-Butyl Hydroperoxide, Menadione, And Neutrophils, Yuh-Cherng Chai, S. Hendrich, James Thomas

Yuh-Cherng Chai

In order to examine potentially important S-thiolated proteins, ^3^5S-labeled hepatocytes were exposed to oxidative stress. A similar group of S-thiolated proteins including carbonic anhydrase III was observed in cells treated with t-butyl hydroperoxide, menadione, or stimulated neutrophils. The radioactive thiols bound to hepatocyte proteins were identified by HPLC and more than 85% was glutathione. In menadione-treated hepatocytes, proteins were gradually S-thiolated over 30 min and 25% of the cellular glutathione pool became protein-bound. In t-butyl hydroperoxide-treated cells, S-thiolation was more transient and 11% of the glutathione was protein-bound. Neutrophil-treated hepatocytes had nearly the same amount of protein S-thiolation (8 ...


S-Thiolation Of Individual Human Neutrophil Proteins Including Actin By Stimulation Of The Respiratory Burst: Evidence Against A Role For Glutathione Disulfide, Yuh-Cherng Chai, S. Ashraf, R. Johnson, James Thomas Mar 1994

S-Thiolation Of Individual Human Neutrophil Proteins Including Actin By Stimulation Of The Respiratory Burst: Evidence Against A Role For Glutathione Disulfide, Yuh-Cherng Chai, S. Ashraf, R. Johnson, James Thomas

Yuh-Cherng Chai

Protein S-thiolation, a reversible modification of protein sulfhydryls resulting in formation of mixed-disulfides, was studied in human neutrophils stimulated with phorbol diester to produce superoxide anion. Rapid S-thiolation of several proteins was examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Glutathione was identified as the primary protein-bound thiol by HPLC chromatography, contributing considerably more than 85% of the total. Minor amounts of homocysteine and/or cysteine were also detected as protein-bound thiols. During the first 30 min after stimulation, 10% of the cellular glutathione became protein bound (2 nmol/mg of protein). There was no increase in glutathione disulfide suggesting that ...


Altering The Rna Binding Specificity Of A Translational Repressor, F Lim, Marc Spingola, D S. Peabody Mar 1994

Altering The Rna Binding Specificity Of A Translational Repressor, F Lim, Marc Spingola, D S. Peabody

Marc Spingola

The coat proteins of RNA phages MS2 and GA are specific RNA-binding proteins which function to encapsidate viral RNA and to translationally repress synthesis of the viral replicase. The two proteins have highly homologous amino acid sequences, yet they show different RNA binding specificities, recognizing RNA stem-loop structures which differ primarily in the nucleotide sequences of their loops. We sought to convert MS2 coat protein to the RNA binding specificity of GA through the introduction of GA-like amino acid substitutions into the MS2 coat protein RNA-binding site. The effects of the mutations were determined by measuring the affinity of the ...


Irreversible Steps In The Ferritin Synthesis Induction Pathway.., L. Linggoess, David Mascotti, M. Bhattacharyya-Pakrasi, H. Gang, R. Thach Feb 1994

Irreversible Steps In The Ferritin Synthesis Induction Pathway.., L. Linggoess, David Mascotti, M. Bhattacharyya-Pakrasi, H. Gang, R. Thach

David P. Mascotti

The ability of cells to re-repress ferritin synthesis after removal of an inducing agent (iron or heme) was investigated. Re-repression was found to be a slow process, requiring approximately 4 (after iron removal) to 10 h (after heme removal) for completion. Desferrioxamine mesylate (Desferal) had only a slight effect on the rate of re-repression, whereas cycloheximide was strongly inhibitory, indicating that new protein synthesis is required for re-repression. Re-repression occurred at a slow but significant rate in the presence of both Desferal and cycloheximide. These results indicate that, in the absence of an iron chelator, the induction of ferritin synthesis ...


Protein S-Thiolation And Dethiolation, James Thomas, Yuh-Cherng Chai, Che-Hun Jung Dec 1993

Protein S-Thiolation And Dethiolation, James Thomas, Yuh-Cherng Chai, Che-Hun Jung

Yuh-Cherng Chai

No abstract provided.


Avian Sleep, Charles Amlaner, Nigel Ball Dec 1993

Avian Sleep, Charles Amlaner, Nigel Ball

Charles J. Amlaner

No abstract provided.


S-Thiolation And Irreversible Oxidation Of Sulfhydryls On Carbonic Anhydrase Iii During Oxidative Stress: A Method For Studying Protein Modification In Intact Cells And Tissues, C. Lii, Yuh-Cherng Chai, W. Zhao, James Thomas, S. Hendrich Dec 1993

S-Thiolation And Irreversible Oxidation Of Sulfhydryls On Carbonic Anhydrase Iii During Oxidative Stress: A Method For Studying Protein Modification In Intact Cells And Tissues, C. Lii, Yuh-Cherng Chai, W. Zhao, James Thomas, S. Hendrich

Yuh-Cherng Chai

No abstract provided.