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Biomedical subjects

J M Cameron

Publications and source records attributed to J M Cameron.

At least 19 recordsLinked to original sources
PubMed2006

Pyruvate dehydrogenase phosphatase deficiency: orphan disease or an under-diagnosed condition?

Pyruvate dehydrogenase phosphatase (PDP) is an enzyme which regulates the activity of the pyruvate dehydrogenase complex (PDHc). In the past, PDHc deficiency has been attributed to mutations in the complex itself and the regulatory enzymes have not been considered. We have recently reported the first mutation in PDP1, one of the two isoforms of PDP, which results in severe exercise intolerance and mild developmental delay in patients. This novel process of aberrant pyruvate metabolism opens up a new avenue for investigation into PDHc deficiency, that has hitherto been underappreciated.

Humans↗
PubMed2005

Computational identification of human mitochondrial proteins based on homology to yeast mitochondrially targeted proteins.

MOTIVATION: Patients with defects of the mitochondrial respiratory chain due to mutations in nuclear genes are often undiagnosable due to the lack of information about the role of these genes. We therefore sought to produce a novel dataset of human nuclear-encoded mitochondrial proteins. RESULTS: We have used the web-based computer program Mitoprot to predict which proteins in the Saccharomyces cerevisiae genome are targeted to mitochondria. We then used this protein dataset to identify the homologous human proteins in the Unigene database using TBLASTN from NCBI. Human proteins with an Expectation value <10(-5) and an Identity >30% were accepted as true homologues of the yeast proteins. These human proteins were then reanalyzed with Mitoprot. The final set of proteins comprises a dataset of 361 human mitochondrially targeted proteins with homology to all S.cerevisiae mitochondrially targeted proteins. One hundred twenty eight of these proteins are novel and are of unknown function. SUPPLEMENTARY INFORMATION: Supplementary tables will be available from http://www.sickkids.ca/Robinsonlab/

Conserved Sequence↗
PubMed2001

TTY2: a multicopy Y-linked gene family.

Genes involved in human male sex determination and spermatogenesis are likely to be located on the Y chromosome. In an effort to identify Y-linked, testis-expressed genes, a cDNA selection library was generated by selecting testis cDNA with Y-cosmid clones. Resultant clones containing repetitive or vector material were eliminated, and 79 of the remaining clones were sequenced. Nineteen cDNAs showed homology with the TTY2 gene, and indicated that TTY2 is part of a large gene family. Screening of a panel of Y-linked cosmids revealed that the TTY2 gene family includes at least 26 members organized in 14 subfamilies. Further investigation revealed that TTY2 genes are arranged in tandemly arrayed clusters on both arms of the Y chromosome, and each gene comprises a series of tandemly arranged repeats. RT-PCR studies for two of these genes revealed that they are expressed in adult and fetal testis, as well as in the adult kidney. None of the genes investigated in detail contain an open reading frame. We conclude that the TTY2 gene family is composed of multiple copies, some of which may function as noncoding RNA transcripts and some may be pseudogenes.

Adult↗
PubMed2000

Use of six chimeric proteins to investigate the role of intramolecular interactions in determining the kinetics of carnitine palmitoyltransferase I isoforms.

The two isoforms of carnitine palmitoyltransferase I (CPT I; muscle (M)- and liver (L)-type) of the mitochondrial outer membrane have distinct kinetic characteristics with respect to their affinity for one of the substrates (l-carnitine) and the inhibitor malonyl-CoA. Moreover, they differ markedly in their hysteretic behavior with respect to malonyl-CoA and in their response to changes in the in vivo metabolic state. However, the two proteins are 62% identical and have the same overall structure. Using liver mitochondria, we have previously shown that the protein is polytopic within the outer membrane, comprising a 46-residue cytosolic N-terminal sequence, two transmembrane segments (TM1 and TM2) separated by a 27-residue loop, and a large catalytic domain (also cytosolic) (Fraser, F., Corstorphine, C. G., and Zammit, V. A. (1997) Biochem. J. 323, 711-718). We have now conducted a systematic study on six chimeric proteins constructed from combinations of three linear segments of rat L- and M-CPT I and on the two parental proteins to elucidate the effects of altered intramolecular interactions on the kinetics of CPT activity. The three segments were (i) the cytosolic N-terminal domain plus TM1, (ii) the loop plus TM2, and (iii) the cytosolic catalytic C-terminal domain. The kinetic properties of the chimeric proteins expressed in Pichia pastoris were studied. We found that alterations in the combinations of the N-terminal plus TM1 and C-terminal domains as well as in the N terminus plus TM1/TM2 pairings resulted in changes in the K(m) values for carnitine and palmitoyl-CoA and the sensitivity to malonyl-CoA of the L-type catalytic domain. The changes in affinity for malonyl-CoA and palmitoyl-CoA occurred independently of changes in the affinity for carnitine. The kinetic characteristics of the M-type catalytic domain and, in particular, its malonyl-CoA sensitivity were much less susceptible to influence by exchange of the other two segments of the protein. The marked difference in the response of the two catalytic domains to changes in the N-terminal domain and TM combinations explains the previously observed differences in the response of L- and M-CPT I to altered physiological state in intact mitochondria and to modulation of altered lipid molecular order of the mitochondrial outer membrane in vivo and in vitro.

Amino Acid Sequence↗
PubMed2000

The state and future of Mars polar science and exploration.

As the planet's principal cold traps, the martian polar regions have accumulated extensive mantles of ice and dust that cover individual areas of approximately 10(6) km2 and total as much as 3-4 km thick. From the scarcity of superposed craters on their surface, these layered deposits are thought to be comparatively young--preserving a record of the seasonal and climatic cycling of atmospheric CO2, H2O, and dust over the past approximately 10(5)-10(8) years. For this reason, the martian polar deposits may serve as a Rosetta Stone for understanding the geologic and climatic history of the planet--documenting variations in insolation (due to quasiperiodic oscillations in the planet's obliquity and orbital elements), volatile mass balance, atmospheric composition, dust storm activity, volcanic eruptions, large impacts, catastrophic floods, solar luminosity, supernovae, and perhaps even a record of microbial life. Beyond their scientific value, the polar regions may soon prove important for another reason--providing a valuable and accessible reservoir of water to support the long-term human exploration of Mars. In this paper we assess the current state of Mars polar research, identify the key questions that motivate the exploration of the polar regions, discuss the extent to which current missions will address these questions, and speculate about what additional capabilities and investigations may be required to address the issues that remain outstanding.

Atmosphere↗
PubMed1999

Sequencing and functional expression of the malonyl-CoA-sensitive carnitine palmitoyltransferase from Drosophila melanogaster.

Using expressed sequence tag data, we obtained a cDNA for a carnitine palmitoyltransferase I (CPT I)-like molecule from Drosophila melanogaster. The cDNA encodes a 782-residue protein that shows 49% and 48% sequence identity with the rat liver and skeletal-muscle isoforms of CPT I respectively. The sequence has two predicted membrane-spanning regions, suggesting that it adopts the same topology as its mammalian counterparts. The sequence contains all the residues that have been shown to be characteristic of carnitine acetyltransferases. Expression in the yeast Pichia pastoris confirmed that the cDNA does encode a CPT enzyme. The activity was found to be associated with a mitochondria-enriched fraction. Kinetic analysis revealed a K(m) for carnitine of 406 microM and a K(m) for palmitoyl-CoA of 105 microM. The CPT activity was very sensitive to inhibition by malonyl-CoA, with an IC(50) of 0.74 microM when the activity was assayed with 35 microM palmitoyl-CoA and 1% (w/v) albumin at pH 7.0. A histidine residue at position 140 in rat liver CPT I has been indicated to be important for inhibition by malonyl-CoA. The equivalent residue (position 136) in Drosophila CPT I is arginine, implying that any basic residue might be compatible with such sensitivity. Evidence is presented that, unlike in mammals, Drosophila has only a single CPT I gene. Sequences suggesting the existence of a splice variant in the 5' untranslated region were found; this was consistent with the existence of two promoters for the CPT I gene.

Amino Acid Sequence↗
PubMed1996

Degeneracy in human multicopy RBM (YRRM), a candidate spermatogenesis gene.

In order to search for mutations in the multicopy RBM genes that might be associated with male infertility, we have used sequence data from the reported cDNA clone to determine the intron exon boundaries of the YRRM 1 gene. This gene has 12 exons, three of which encode the putative RNA binding domain of the protein. Different copies of the gene contain sequence variations and, additionally, give rise to transcripts with different numbers of copies of the repeated SRGY motif. Since mutations in the RNA binding domain would seem likely to have an effect on the activity of the protein, we have scanned these exons for mutations by SSCP on DNA from normal and infertile men. Sequence differences in the exon encoding the N-terminal part of the RNA binding domain account for at least four different classes of the gene and give rise to different SSCP conformers. Sequence analysis shows that one of these classes is a pseudogene and that the members of another class are nonfunctional. RT-PCR shows that all classes are transcribed and that the A class is most abundant. We have found a point mutation that alters the highly conserved RNP2 motif in one infertile patient. This mutation is also found in his father. We have used PCR followed by SSCP analysis to map RBM on a Y Chromosome (Chr) YAC contig and have demonstrated a distribution that spans a major part of this chromosome's euchromatin.

Amino Acid Sequence↗
PubMed1995

The intracellular phosphorylation of (-)-2'-deoxy-3'-thiacytidine (3TC) and the incorporation of 3TC 5'-monophosphate into DNA by HIV-1 reverse transcriptase and human DNA polymerase gamma.

(-)-2'-deoxy-3'-thiacytidine (3TC) has been shown to be a potent, selective inhibitor of HIV replication in vitro, which requires phosphorylation to its 5'-triphosphate for antiviral activity. The intracellular concentration of 3TC 5'-triphosphate in phytohaemagglutinin (PHA)-stimulated peripheral blood lymphocytes (PBL) shows a linear dependence on the extracellular concentration of 3TC up to an extracellular 3TC concentration of 10 microM. At this extracellular concentration of 3TC, the resulting intracellular concentration of 3TC 5'-triphosphate is 5 microM. This value is similar to the inhibition constant (Ki) values for the competitive inhibition of human immunodeficiency virus type 1 (HIV-1) reverse transcriptase and human DNA polymerases (10-16 microM) by 3TC 5'-triphosphate. Since the concentration of 3TC producing 90% inhibition (IC90) of HIV replication in PBLs has been reported to be 76 nM, the antiviral activity of 3TC requires intracellular concentrations of 3TC 5'-triphosphate, which would result in very little inhibition of reverse transcriptase if its sole mode of action was competitive inhibition. This apparent discrepency may be explained by the ability of 3TC 5'-triphosphate to act as a substrate for reverse transcriptase. Primer extension assays have shown that 3TC 5'-triphosphate is a substrate for HIV-1 reverse transcriptase and DNA polymerase gamma, resulting in the incorporation of 3TC 5'-monophosphate into DNA. In the case of DNA polymerase gamma, the product of this reaction (i.e. double-stranded DNA with 3TC 5'-monophosphate incorporated at the 3'-terminus of the primer strand) is also a substrate for the 3'-5' exonuclease activity of this enzyme. This may explain the low levels of mitochondrial toxicity observed with 3TC.

Base Sequence↗
PubMed1995

Atypical migratory stomatitis and Munchausen syndrome presenting as periorbital ecchymosis and mandibular subluxation.

Stomatitis areata migrans, unlike its analogue on the tongue, migratory glossitis, is not easily recognized and is so uncommon and varied in appearance that it may escape definitive diagnosis. It may be so puzzling to the clinician that the patient's credibility may be questioned. A detailed report of a case is presented in which an atypical migratory stomatitis went undiagnosed. Bizarre patient behavior followed in the form of self-inflicted injury (Munchausen syndrome) as the patient attempted to convince the care providers of the true existence of lesions in order to maintain their interest and to obtain relief from discomfort.

Adult↗
PubMed1994

Discovery and analysis of a series of C2-symmetric HIV-1 proteinase inhibitors derived from penicillin.

In order to identify a suitable peptide substrate for human immunodeficiency virus-1 (HIV-1) proteinase, a range of peptides from various cleavage sites within the gag-pol polyprotein were assayed by HPLC for specific cleavage. The peptide with the optimal combination of favorable kinetics and good solubility was based on the N-terminus cleavage site of HIV-1 proteinase (KQGTVSFNF*PQIT). The HPLC assay, using the above peptide, was developed into a rapid isocratic method in order to analyze inhibition kinetics. An assay suitable for high-throughput screening was developed using a radioactively labeled peptide with the same sequence, coupled to a solid phase. Using this assay, a C2-symmetric HIV-1 proteinase inhibitor derived from penicillin was discovered during random screening of a compound library. A chemical synthesis program developed this structure into a series of potent inhibitors. The lead structures were highly selective for HIV-1 proteinase with good antiviral activity in vitro against HIV and no cytotoxicity. The HPLC assay was used to demonstrate that these compounds are competitive tight-binding inhibitors of HIV-1 proteinase.

Amino Acid Sequence↗
PubMed1994

Susceptibility in cell culture of feline immunodeficiency virus to eighteen antiviral agents.

The in-vitro susceptibilities of two strains of feline immunodeficiency virus to 18 antiviral agents were determined in two cell lines. In terms of inhibiting p24 antigen production, the nucleoside-analogue reverse transcriptase inhibitors were the most effective compounds. Inhibition was also observed with aurintricarboxylic acid, phosphonoformate and butyldeoxynorjirimycin, but not with the other agents tested.

Animals↗
PubMed1993

Royal College of Surgeons comparative ENT audit 1990.

The Royal College of Surgeons Comparative Audit Service was set up in 1990 so that surgeons could pool their audit data, to provide 'standards' with which to compare their own figures. A total of 405 consultant otolaryngologists were circularized in December 1991 inviting them to return data about their resources, workload, case-mix and complications, and about two specific audit topics-Myringoplasty and Carcinoma of the Larynx--for the calendar year 1990. A total of 65 consultants returned proformas with data on 52208 admissions and 31240 surgical procedures. The 'average' respondent admitted 829 patients in the year (19% of these day cases and 14% emergencies) and performed 744 surgical procedures with a mean complication rate of 1.39% using three theatre sessions per week. Cancelled theatre sessions per annum per consultant ranged from 0 to 71. The mean known success rate after myringoplasty was 65%, with hearing improvement in 53%. The 'average' ENT surgeon saw 3.5 new cases of invasive carcinoma of the larynx and treated 69% of these with radiotherapy alone, compared with 14% surgery alone. As well as allowing a profile to be drawn up of the 'average' respondent, this audit allowed individual consultants returning data to compare their own figures in detail with the pooled data, which were presented graphically at a meeting in April 1992.

Carcinoma↗
PubMed1993

High-level resistance to (-) enantiomeric 2'-deoxy-3'-thiacytidine in vitro is due to one amino acid substitution in the catalytic site of human immunodeficiency virus type 1 reverse transcriptase.

Passage of human immunodeficiency virus type 1 in the presence of increasing 2'-deoxy-3'-thiacytidine (3TC) concentrations results in high-level (> 100-fold) 3TC-resistant viruses. All 3TC-resistant viruses possess a substitution at the second codon (from a methionine into an isoleucine) at position 184 within the highly conserved motif (YMDD) of human immunodeficiency virus type 1 reverse transcriptase. 3TC-resistant viruses were cross-resistant to the (-) enantiomer of the fluorinated derivative of BCH-189 but remained susceptible to 2',3'-dideoxyinosine and 2',3'-dideoxycytidine. The susceptibilities of the 3TC-resistant viruses to the (+) enantiomers of BCH-189 and the fluorinated derivative of BCH-189 demonstrate an enantiomeric specificity for viruses selected under these conditions. Introduction of an isoleucine substitution at codon 184 into a background of two known 3'-azido-3'-deoxythymidine resistance mutations (amino acids 41 and 215) restored the susceptibility of this virus to 3'-azido-3'-deoxythymidine.

Antiviral Agents↗
PubMed1993

Plastic bag suffocation.

Suffocation by plastic bag is not common. The finding of a body at the scene of death with a plastic bag in situ could be diagnostically valuable to the pathologist. Otherwise the pathologist may be hard put to arrive at a firm opinion as to how the deceased met his/her death; in fact, in three of the cases reported in this paper, the local pathologist could not give a cause of death when the initial autopsy was performed. Plastic bags could be accidentally placed over the head; put there to help in the inhalation of volatile substances; with homicidal intention; or simply as an effective suicidal method. The object of this paper is to study the cases where bodies were found with a plastic bag over the head, and in particular the cases where the cause of death was due to plastic bag suffocation.

Adult↗
PubMed1993

The diagnosis of drowning by quantitative and qualitative diatom analysis.

The reliability and applicability of quantitative and qualitative diatom analysis in the diagnosis of drowning has been evaluated. Water and organ samples of immersion cases reported in the area covered by the Department of Forensic Medicine of the London Hospital Medical College, were analysed using both light and scanning electron microscopy. Controls included organ samples of the bodies of people who died from natural causes and exclusion of contamination. Organ samples of both immersion and control cases were prepared by chemical digestion with concentrated nitric acid. Diatoms were present in the majority of samples of organs of both immersion and control cases but there was a significant quantitative difference between the number of diatoms in control and immersion cases. Qualitative analysis of water and organ samples of immersion cases supported the diagnosis of death due to aspiration of water in approximately a third of the total of bodies found in water. It has been suggested that the present analysis can be used as basic criteria for standardization of the diatom method.

Brain↗