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Biomedical subjects

G R Sutherland

Publications and source records attributed to G R Sutherland.

At least 181 records · Page 10Linked to original sources
PubMed1996

The effects of calcium on the thermal stability and activity of manganese peroxidase.

The presence of micromolar Ca2+ efficiently prevented the thermal inactivation of manganese peroxidase from Phanerochaete chrysosporium. The amount of Ca2+ normally present in the enzyme decreased when the enzyme was thermally inactivated and EGTA increased the rate of inactivation. The inactivation kinetics were biphasic, suggesting a sequential two-step process. The rate of inactivation during the second, slower step corresponded to the rate of loss of heme from the enzyme. Thermally inactivated manganese peroxidase could be readily reactivated in the presence of excess Ca2+. However, as the time of thermal incubation increased and the amount of remaining heme decreased, the amount of enzyme activity recovered decreased. Therefore, while both steps of denaturation could be prevented by Ca2+, only one step could be reversed upon the addition of Ca2+. It is proposed that the first step of denaturation involves the loss of Ca2+ which causes conformational changes resulting in the loss of manganese peroxidase activity. The second step is believed to involve further structural loss and results in the loss of heme from the enzyme. It is concluded that manganese peroxidase is susceptible to thermal inactivation because it contains relatively labile Ca2+ ions required for stability and activity.

Basidiomycota↗
PubMed1996

Structure and chromosomal localization of the genomic locus encoding the Kiz1 LIM-kinase gene.

We have cloned and characterized the mouse gene encoding Kiz1/Limk1, a new member of the zinc-finger LIM family that also has a kinase domain. The gene encompasses 25 kb of the mouse genome, and the organization of its 16 exons does not correlate with its functional domains. The promoter region of Kiz1/Limk1 was identified by cloning a 1.06-kb genomic fragment upstream from the first ATG in a promotorless CAT vector. This construct was demonstrated to drive CAT expression in Jurkat cells. The promoter sequence lacks conventional TATA and CAAT motifs but contains consensus binding sequences for several transcriptional regulators implicated in control of transcription in many different cell types, including Sp1, Ets, and E2A. Analysis of the chromosomal localization of KIZ1/LIMK1 indicates that it lies on human chromosome 17 in the region 17q25 and on mouse Chromosome 5, band G2.

Animals↗
PubMed1996

Characterization and chromosomal localization of the human A2a adenosine receptor gene: ADORA2A.

The gene for the stimulatory G protein-coupled human A2a adenosine receptor was isolated and sequence analysis revealed two exons that are interrupted by an intron of approximately 6.4 kb. An intron is located in the same region in the human A1 and A2b adenosine receptor genes. Comparison of the A2a genomic and cDNA sequences reveals two nucleotide differences in the coding region and the presence of an aberrant sequence in the 5'208 base pairs of the A2a cDNA including a polymorphism in the third base of codon Tyr-361 and Gly codon which was always detected at residue 392, indicated that the Arg codon present in the cDNA may be an artifact. Fluorescent in situ hybridization and PCR analysis of human-hamster hybrid cell panels shows that the A2a receptor gene is localized to chromosome 22q11.2. This is in contrast with previous reports (subsequently retracted) which mapped the A2a gene to chromosome 11q11-13.

Amino Acid Sequence↗
PubMed1996

The effect of veratryl alcohol on manganese oxidation by lignin peroxidase.

The extracellular peroxidase isozymes secreted by the white rot fungus Phanerochaete chrysosporium have been classified as manganese peroxidases (isozymes H3, H4, H5, and H9) and lignin peroxidases (isozymes H1, H2, H6, H7, H8, and H10). Recently we reported peroxidase isozyme H2 can also oxidize Mn2+ (Khindaria et al., 1995, Biochemistry 34, 7773-7779). This lignin peroxidase isozyme oxidized Mn2+ with both of the enzyme intermediates, compound I and compound II, at the same rates as manganese peroxidase isozyme H4. The results of single-turnover kinetic studies have now demonstrated that compound I of the other lignin peroxidase isozymes (H1, H6, H7, H8, and H1O) also readily oxidized Mn2+, but that the rate of Mn2+ oxidation by compound II was extremely slow. Compound III rapidly the presence of Mn2+, oxalate, and H2O2. However, upon the addition of veratryl alcohol, the results indicated that veratryl alcohol served to reduce compound II. Under such conditions, compound III did not accumulate, and a steady-state rate of Mn2+ oxidation was observed. The rate of Mn2+ oxidation was the same as for the reduction of compound II by veratryl alcohol. The dependence of the rate of Mn2+ oxidation on the concentration of veratryl alcohol was consistent with a mechanism in which Mn2+ is oxidized by compound I and veratryl oxidized by compound II. Therefore, under physiologically relevant conditions, in which both veratryl alcohol and Mn2+ are present, all lignin peroxidase isozymes would be capable of oxidizing Mn2+ to Mn3+ which can serve as a diffusible oxidant.

Agaricales↗
PubMed1996

Further characterization of proteins associated with elastic fiber microfibrils including the molecular cloning of MAGP-2 (MP25)

Together with the 31-kDa microfibril-associated glycoprotein (MAGP), four polypeptides designated MP340 (340 kDa), MP78 (78 kDa), MP70 (70 kDa), and MP25 (25 kDa) have previously been identified in tissue extracts designed specifically to solubilize the microfibrillar component of elastic fibers. In the present study, both MP78 and MP70 were shown to be forms of a protein which is closely related to the human protein beta ig-h3, and MP340 was confirmed to be the bovine form of fibrillin-1. Peptide sequences from MP25 proved to be unique, and affinity-purified anti-MP25 antibodies were shown, by immunofluorescence and immunoelectron microscopy, to localize specifically to the elastin-associated microfibrils. This confirmed that MP25 was a distinct component of these structures. Expression screening of nuchal ligament cDNA libraries yielded a cDNA, cM10A (770 base pairs) which encodes amino acid sequences matching those of the MP25 peptides. Further library screening with cM10A identified cDNAs which encode the complete primary structures of bovine and human MP25. Bovine and human MP25 were found to be around 80% homologous and contain 170 and 173 amino acids, respectively. Data base searches revealed that MP25 had significant similarity of structure only with MAGP, indicating that the two proteins form a new family of microfibrillar proteins. In acknowledgment, MP25 has been formally renamed MAGP-2, and MAGP is referred to as MAGP-1. The close similarity between the two proteins (57%) is confined to a central region of 60 amino acids where there is precise alignment of 7 cysteine residues. Elsewhere the MAGP-2 molecule is rich in serine and threonine residues and contains an RGD motif. MAGP-2 lacks the proline-, glutamine-, and tyrosine-rich sequences and a hydrophobic carboxyl terminus, characteristic of MAGP-1. These structural differences suggest that MAGP-2 has some functions which are distinct from those of MAGP-1. The locus of the human MAGP-2 gene was identified on chromosome 12 in the region of 12p12.3-12p13.1.

Amino Acid Sequence↗
PubMed1996

Fragile X syndrome and fragile XE mental retardation.

There are two forms of mental handicap associated with fragile sites on the end of the long arm of the X chromosome. The well known common disorder Fragile X syndrome is associated with FRAXA and a rare non-specific form of mental handicap is associated with FRAXE. The cytogenetics of these fragile sites is considered. For Fragile X syndrome details are given of the molecular genetics, inheritance patterns, genetic counselling, methods for diagnosis of index cases, carrier detection and prenatal diagnosis. Series of prenatal diagnoses are briefly reviewed and technical and biological problems associated with this procedure are considered. Prenatal diagnosis of Fragile X syndrome using molecular genetic techniques is now a well established procedure, with the only significant problem being the inability to accurately predict phenotype in female fetuses with full mutations. Few prenatal diagnoses of Fragile XE non-specific mental retardation have been recorded. In principle the technical aspects of such a prenatal diagnosis should be little different from those for Fragile X syndrome. Incomplete knowledge of the phenotypic effect of the full mutation in males and females would make phenotypic prediction for any fetus shown to have such a mutation very difficult. At this stage all that could be determined with precision is that the mutation was present or absent in the fetus. Possible consequences of this are discussed.

Female↗
PubMed1996

Unusual inheritance patterns due to dynamic mutation in fragile X syndrome.

Fragile X syndrome is the most common form of familial mental retardation and is one of the world's most common genetic diseases. The inheritance patterns of the disease have many unusual features. It is an X-linked disorder yet there are asymptomatic carrier males. The disease is expressed only when the gene is inherited from the mother. The risk of a carrier woman having a child with the syndrome depends upon her position in the pedigree (the Sherman paradox) and her own intellectual status. The discovery that the disease is due to dynamic mutation (which is a multistage process) that inactivates FMR1 has provided an explanation for the unusual inheritance patterns. The finding of linkage disequilibrium between the fragile X mutations and closely linked DNA markers (haplotype) has required a reinterpretation of this phenomenon for dynamic mutations. Only a small number of normal alleles at the fragile X locus have long stretches of perfect repeat (2% with more than 24 copies) and these form a reservoir of alleles that can increase in length into the premutation range. Dynamic mutation is, so far, an exclusively human phenomenon, but this is probably because it has yet to be discovered in other species. Unusual inheritance patterns are a hallmark of dynamic mutation diseases.

Alleles↗
PubMed1996

Isolation and characterization of cDNA clones for Humly9: the human homologue of mouse Ly9.

Ly9 is a mouse cell membrane antigen found on all lymphocytes and coded for by a gene that maps to chromosome 1. We previously described the isolation and characterization of a full-length cDNA clone for mouse Ly9. Using cross-species hybridization we isolated cDNA clones encoding the human homologue Humly9. Analysis of the predicted protein sequence suggests that the extra-cellular portion of the Humly9 molecules is composed of four Ig-like domains: a V domain (V) without disulphide bonds and a truncated C2 domain (tC2) with two disulphide bonds, a second V domain without disulphide bonds and a second tC2 with two disulphide bonds, i.e., as V-tC2-V-tC2. The gene encoding Humly9 was mapped to chromosome 1 by analysis of human/hamster hybrids, and more specifically to the 1q22 region by in situ hybridization. The protein sequence data support the view that Humly9 belongs to the immunoglobulin-superfamily subgroup which includes CD48, CD2, and LFA-3.

Amino Acid Sequence↗
PubMed1996

Asymptomatic large pituitary adenomas discovered at autopsy.

BACKGROUND: Pituitary adenomas over 1 inch (2.5 cm) in diameter might seem to warrant surgical intervention by virtue of their inevitable compression and distortion of surrounding neural tissues. METHODS: We report three pituitary adenomas over 1 inch in diameter, causing gross distortion of the optic nerves, chiasm, and tracts, as well as surrounding brain structures. All three tumors were asymptomatic and were discovered incidentally at autopsy in patients 65, 79, and 101 years of age. RESULTS: Tumor diameters were 2.5, 4.0, and 2.5 cm, respectively. Histologically, all were chromophobe adenomas, free of either hemorrhage or necrosis. In the absence of hormonal secretion or pituitary apoplexy, symptom-free giant adenomas of the pituitary are compatible with a long life. CONCLUSIONS: We conclude that the need for surgical intervention in pituitary adenomata should be determined by considerations other than mere tumor presence and size.

Adenoma↗
PubMed1996

Verification of cardiac doppler tissue images using grey-scale M-mode images.

Doppler tissue imaging (DTI) is a new ultrasonic technique that can be used to give quantified information about the motion of the myocardium. DTI M-mode recordings can be generated that display cardiac wall motion in great detail. In order to verify the motion as displayed in these images comparisons were made with simultaneously obtained grey-scale M-mode recordings. After capturing 135 simultaneous DTI and grey-scale M-mode recordings, those were selected in which wall motion could be accurately assessed from the grey-scale recording. Comparison with the DTI images shows: (1) that DTI accurately displays the direction of wall motion; and (2) that DTI displays whether the wall is thickening or thinning as a velocity distribution across the heart wall. This information is more reliably displayed and easier to interpret in the DTI M-mode recordings than in the grey-scale M-mode recordings.

Animals↗
PubMed1996

DNAM-1, a novel adhesion molecule involved in the cytolytic function of T lymphocytes.

Intercellular adhesion molecules play an important role in the generation of T lymphocyte-mediated immune responses. Here, we describe a novel accessory molecule, DNAX accessory molecule-1 (DNAM-1), that is constitutively expressed on the majority of peripheral blood T lymphocytes. DNAM-1 is a 65 kDa transmembrane glycoprotein consisting of 318 aa including two immunoglobulin-like domains. Anti-DNAM-1 monoclonal antibody (MAb) inhibits T and NK cell-mediated cytotoxicity against a variety of tumor cell targets and blocks cytokine production by alloantigen-specific T cells. In addition, DNAM-1 is a tyrosine-phosphorylated signal-transducing molecule that participates in primary adhesion during cytotoxic T lymphocyte (CTL)-mediated cytotoxicity.

Amino Acid Sequence↗
PubMed1996

Epidemiology of childhood brain tumors.

BACKGROUND: Brain tumors comprise more than 20% of all childhood malignancies, and constitute the greatest number of solid pediatric cancers. Incidence rates reported have varied from 2.4 to 3.5/100,000 children, reflecting the impact of modern imaging techniques, the application of diverse investigative methodologies, and the accessibility of the community to health care. METHODS: Material from patients < 18 years of age was collated from the Manitoba Cancer Foundation Tumor Registry, the personal records of Winnipeg pediatric neurologists, and autopsy data. Patient data were also obtained from hospital charts and operating room log books. Histological sections were examined and classified according to the American Cancer Society by a single neuropathologist. The chi-square test was used for statistical evaluation. RESULTS: During the seven-year study period, the diagnosis of brain tumor was made in 89 pediatric patients, of which 88 were diagnosed premortem. The overall average annual incidence rate for both sexes was 4.03/100,000 child-years, higher than that previously reported. The male and female average annual incidence rates were 4.2 and 3.7/100,000 child-years, respectively. Tumor type and location were relatively unremarkable, with an expected peak of medulloblastoma occurring in young males. The yearly incidence of tumor occurrence was fairly stable, and the geographic distribution of cases within Manitoba, homogeneous. CONCLUSION: The highest incidence rates of pediatric brain tumors have been recorded in countries possessing sophisticated universal health care systems, possibly reflecting their efficacy in disease surveillance.

Adolescent↗
PubMed1996

Identification of the gene FMR2, associated with FRAXE mental retardation.

Five folate-sensitive fragile sites have been characterized at the molecular level (FRAXA, FRAXE, FRAXF, FRA16A and FRA11B). Three of them (FRAXA, FRAXE and FRA11B) are associated with clinical problems, and two of the genes (FMR1 in FRAXA and CBL2 in FRA11B) have been identified. All of these fragile sites are associated with (CCG)n/(CGG)n triplet expansions which are hypermethylated beyond a critical size. FRAXE is a rare folate sensitive fragile site only recently recognized. Its cytogenetic expression was found to involve the amplification of a (CCG)n repeat adjacent to a CpG island. Normal alleles vary from 6 to 25 copies. Expansions of greater than 200 copies were found in FRAXE expressing males and their FRAXE associated CpG island was fully methylated. An association of FRAXE expression with concurrent methylation of the CpG island and mild non-specific mental handicap in males has been reported by several groups. We now report the cloning and characterization of a gene (FMR2) adjacent to FRAXE. Elements of FMR2 were initially identified from sequences deleted from a developmentally delayed boy. We correlate loss of FMR2 expression with (CCG)n expansion at FRAXE, demonstrating that this is a gene associated with the CpG island adjacent to FRAXE and contributes for FRAXE-associated mild mental retardation.

Amino Acid Sequence↗
PubMed1996

Regional variations of ultrasonic integrated backscatter in normal and myopathic left ventricles. A new multi-view approach.

The purpose of the present study was to determine whether the cyclic variation of integrated backscatter is measurable and quantifiable in all left ventricular walls and whether the information obtained using both parasternal and apical transducer positions can be used to identify changes in myocardial structure and contractility. The cyclic variation of integrated backscatter was measured from the parasternal long-axis, apical four-chamber and two-chamber views in 26 patients with idiopathic dilated cardiomyopathy (mean age 58 +/- 9 years; ejection fraction 29 +/- 10%) and compared with information obtained from 30 aged-matched healthy volunteers. For each subject, the cyclic variation of integrated backscatter was calculated from 16 predetermined regions-of-interest located within the myocardium of the basal and mid-segments of the left ventricle imaged from the long-axis view and also the basal mid and apical left ventricular segments imaged from the two apical views. The cyclic variation of integrated backscatter was found to be present in 100% of the analysed regions-of-interest in healthy volunteers and in 87.5% of the analysed regions-of-interest in patients with idiopathic dilated cardiomyopathy. The mean value of cyclic variation of integrated backscatter, averaged from all regions-of-interest in the idiopathic dilated cardiomyopathy group, was significantly reduced compared to that in the healthy volunteers group (3.2 +/- 2.5 dB [mean +/- SD] vs 4.8 +/- 2.9 dB, P < 0.0001). Additionally, the healthy volunteers group demonstrated marked regional variability in the magnitude of cyclic variation of integrated backscatter which closely followed the regional changes in the contractile function of the normal heart. These regional differences in the magnitude of the cyclic variation of integrated backscatter were only partially retained in the idiopathic dilated cardiomyopathy group, and suggest that a multi-view approach of the recording of cyclic variation of integrated backscatter can be of value to differentiate normal from myopathic myocardium and to quantify regional differences in myocardial contractile performance throughout the left ventricular walls.

Adult↗