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Biomedical subjects

C P Gray

Publications and source records attributed to C P Gray.

12 recordsLinked to original sources
PubMed1998

Strategies towards a better understanding of antibiotic action: folate pathway inhibition in Haemophilus influenzae as an example.

Two-dimensional electrophoresis was applied to the global analysis of the cellular response of Haemophilus influenzae to sulfamethoxazole and trimethoprim, both inhibitors of tetrahydrofolate synthesis. Deregulation of the synthesis rate of 118 proteins, involved in different metabolic pathways, was observed. The regulation of the genes involved in the metabolism of the amino acids methionine, threonine, serine, glycine, and aspartate was investigated in detail by analysis of protein synthesis and Northern hybridization. The results suggested that the synthesis of methionine biosynthetic enzymes in H. influenzae is regulated in a similar fashion as in Escherichia coli. A good correlation between the results obtained by Northern hybridization and quantification of protein synthesis was observed. In contrast to trimethoprim, sulfamethoxazole triggered the increased synthesis of the heat shock proteins DnaK, GroEL, and GroES.

5-Methyltetrahydrofolate-Homocysteine S-Methyltran↗
PubMed1997

Antibody responses and protective immunity to recombinant vaccinia virus-expressed bluetongue virus antigens.

The role of individual viral proteins in the immune response to bluetongue virus (BTV) is not clearly understood. To investigate the contributions of the outer capsid proteins, VP2 and VP5, and possible interactions between them, these proteins were expressed from recombinant vaccinia viruses either as individual proteins or together in double recombinants, or with the core protein VP7 in a triple recombinant. Comparison of the immunogenicity of the vaccinia expressed proteins with BTV expressed proteins was carried out by inoculation of rabbits and sheep. Each of the recombinants was capable of stimulating an anti-BTV antibody response, although there was a wide range in the level of response between animals and species. Vaccinia-expressed VP2 was poorly immunogenic, particularly in rabbits. VP5, on the whole, stimulated higher ELISA titers in rabbits and sheep and in some animals in both species was able to stimulate virus neutralizing antibodies. When the protective efficacy of VP2 and VP5 was tested in sheep, vaccinia-expressed VP2, VP5 and VP2 + VP5 were protective, with the most consistent protection being in groups immunized with both proteins.

Animals↗
PubMed1984

Sequence of the putative origin of replication in the UL region of herpes simplex virus type 1 ANG DNA.

Interest has been stimulated concerning the region mapping between 0.38 and 0.42 on the prototype configuration of the herpes simplex virus type 1 (HSV-1) genome due to the high probability of the presence there of a second origin of DNA replication. A 960 bp restriction fragment (HinfI E) of a class II defective HSV-1 ANG DNA has been sequenced using the viral DNA rather than molecularly cloned DNA. This fragment includes the BamHI U/R cleavage site, mapping at approximately 0.4. Part of the sequence derived in this study displays homology with the origins of DNA replication contained in TRS/IRS of HSV-1 and HSV-2 DNA. The homologous region comprising 76 bp occurs as two copies, each of which contains two palindromically arranged copies of an 8 bp sequence identical to the 'consensus' sequence reported to be part of the origin of DNA replication at the terminus of the mammalian adenoviruses. It can be deduced from a comparison of this structure to the TRS/IRS origin of HSV-1 and HSV-2 that there are two origins of replication in the UL region of HSV-1 ANG DNA. Assuming that the orientation of the consensus sequence is relevant to the direction of DNA replication, one can conclude that the UL origin(s) of HSV-1 ANG is (are) bidirectional. It has not yet been possible to clone DNA fragments molecularly which include the region spanning the UL origin(s) of HSV-1 DNA.

Base Sequence↗
PubMed1984

A study of interfering herpes simplex virus DNA preparations containing defective genomes of either class I or II and the identification of minimal requirements for interference.

Progeny DNA of herpes simplex virus type 1 (HSV-1) strain ANG from infections involving defective interfering virus particles (DI DNA) has been described to be of low infectivity in transfection assays due to the presence of viral genomes that interfere with plaque formation by infectious standard genomes. In this study it is shown that this observation applies both for DI DNA containing repetitive defective DNA of the class I type and for DI DNA containing class II-type defective DNA. Restriction endonucleases with recognition sites only in one class of repetitive defective DNA could be used to reduce selectively the interfering activity of DI DNA preparations containing the respective defective DNA in abundance. The results obtained directly implicate repetitive defective DNA as an interfering agent. Restriction endonucleases that create monomeric DNA fragments from class II HSV-1 ANG defective DNA did not abolish the interfering activity of DI DNA containing this type of defective DNA in high abundance, indicating that it is not simply the repetitive nature of defective DNA that is required for interference. Certain DNA fragments shorter than the repeat unit of repetitive defective DNA were still capable of causing interference even in the absence of cohesive single-stranded ends. The common location of cis recognition signals responsible for progeny DNA maturation and initiation of DNA replication on one DNA fragment, however, appeared to be a minimal requirement for interference by fragmented defective DNA.

Animals↗
PubMed1981

Amplification of a short nucleotide sequence in the repeat units of defective herpes simplex virus type 1 Angelotti DNA.

It has been shown earlier that the reiterated regions TRS and IRS bracketing the Us segment of herpes simplex virus type 1 Angelotti DNA are heterogeneous in size by stepwise insertion of one to six copies of a 550-base-pair nucleotide sequence. Considerably higher amplification of this sequence was observed in defective viral DNA: up to 14 copies were detected to be inserted in the repeat units of a major class of defective herpes simplex virus type 1 Angelotti DNA, dDNA1, which originated from noncontiguous sites located in UL and the inverted repeats of the S component of the parental genome. Physical maps were established for the cleavage sites of KpnI, PstI, XhoI, and BamHI restriction endonucleases on the repeats of dDNA1. The map position of the insertion sequence was determined. It was demonstrated that the amplified inserts were not distributed at random among or within the repeats. A given total population of dDNA1 molecules consisted of different homopolymers, each of which contained a constant number of inserts in all of its repeats. Assuming that a rolling-circle mechanism is involved in the generation of full-length defective herpes simplex virus type 1 Angelotti DNA from single repeat units, these data suggest that the 550-base-pair sequence is amplified in the repeats before the replication process.

DNA Replication↗
PubMed1979

Partial primary structure of bacteriorhodopsin: sequencing methods for membrane proteins.

The sequence of 102 amino acid residues from the NH2 terminus and that of 39 amino acid residues from the COOH terminus of bacteriorhodopsin have been determined. These results are in agreement with those recently published by Ovchinnikov and coworkers [Ovchinnikov, Y.A., Abdulaey, N.G., Feigina, M.Y., Kiselev, A.V. & Lobanov, N.A. (1977) FEBS Lett. 84, 1-4]. Chymotryptic cleavage of bacteriorhodopsin produced two fragments, C-1 (Mr 19,000) and C-2 (Mr 6900), the latter containing the blocked NH2 terminus (pyroglutamic acid). Further fragmentation with CNBr gave mostly hydrophobic fragments, which were separated by gel permeation and reverse-phase high-pressure liquid chromatography in formic acid/ethanol/water mixtures. The fragments were sequenced by a judicious combination of mass spectrometric peptide sequencing and automated Edman degradation. The C-2 fragments were ordered on the basis of methionine-containing peptides identified by gas chromatographic mass spectrometry, while C-1 and C-2 were arranged by analysis of an overlapping CNBr fragment.

Amino Acid Sequence↗
PubMed1979

Amino acid sequence of bacteriorhodopsin.

The complete primary structure of the purple membrane protein bacteriorhodopsin, which contains 248 amino acid residues, has been determined. Methods used for separation of the hydrophobic fragments included gel permeation and reverse-phase high-pressure liquid chromatography in organic solvents. The amino acid sequence was determined by a combination of automatic Edman degradation and mass spectrometric methods. The total sequence was derived by ordering of the CNBr fragments on the basis of methionine-containing peptides identified by gas chromatographic mass spectrometry and by analysis of N-bromosuccinimide fragments containing overlaps between CNBr fragments. The present sequence differs from that recently reported by Ovchinnikov and coworkers with respect to an additional tryptophan (position 138) and several amino acid assignments.

Amino Acid Sequence↗
PubMed1978

Structure of the orgin of DNA replication of bacteriophage fd.

An RNA-polymerase-protected DNA fragment of 125 nucleotides from the origin of single-strand to double-strand replication of bacteriophage fd (ori-DNA) was located on the physical map of the phage genome. A stretch of 187 base pairs of DNA including the ori-DNA was sequenced. This DNA segment contains regions with a highly asymmetric pyrimidine/purine distribution next to regions with 2-fold symmetry that form stable hairpin structures in the viral DNA strand.

Base Sequence↗
PubMed1977

Orientation of bacteriorhodopsin in Halobacterium halobium as studied by selective proteolysis.

The orientation of bacteriorhodopsin in the purple membrane of Halobacterium halobium has been studied by proteolytic degradation of purple membrane sheets, reconstituted vesicles, and whole cells, with the following results: (i) Bacteriorhodopsin in purple membrane sheets is cleaved at a single site by Pronase or trypsin; a polypeptide segment of about 15 amino acids is lost from the carboxyl end. Carboxypeptidase A sequentially releases amino acids from the carboxyl end; the tetrapeptide sequence -Ala-Ala-Thr-Ser(COOH) was tentatively deduced for this terminus. (ii) The apomembrane, which lacks retinal, undergoes a second cleavage with trypsin releasing a fragment of approximately 6300 molecular weight from the amino terminus. (iii) Vesicles reconstituted from the purple membrane sheets and synthetic lecithins, in which the direction of proton pumping is opposite to that in the whole cells, have the carboxyl terminus of bacteriorhodopsin accessible to proteolysis. (iv) In envelope vesicles, which largely pump protons in the same direction as the whole cells, the carboxyl terminus is largely protected against proteolysis. (v) Treatment of whole cells with proteinase K hydrolyzes the cell wall proteins but has no effect on acteriorhodopsin. However, the same treatment after lysis of the cells results in degradation of the hydrophilic region at the carboxyl terminus. The results show that the carboxyl terminus as well as the additional cleavage site near the amino terminus observed in apomembrane are on the cytoplasmic side of the purple membrane.

Apoproteins↗
PubMed1976

Reticulum cell sarcoma of testes.

Seven cases of primary reticulum cell sarcoma of the testes have been added to the world literature, bringing the total to 83 cases. Primary reticulum cell sarcoma should be suspected in all patients over the age of fifty years who have a testicular mass. There is a marked tendency of bilateral involvement of this tumor, as well as an association with cutaneous and pharyngeal lesions. Although there have been a few long-term survivors, over-all prognosis is poor.

Adult↗