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1.
The reactions of purified, homogeneous bovine testicular hyaluronidase have been studied with radioactively labeled oligomers of hyalobiuronic acid, (GlcUA-GlcNAc)n, as substrates and acceptors. Transglycosylation occurs by transfer of a glycosyl residue with retention of configuration from a leaving group to an acceptor. On the basis of detailed examination of cleavage and transglycosylation patterns for the trimer; comparison of trimer, tetramer, and polymer as substrates; comparison of acceptors; equilibrium binding; and other data, it is proposed that the enzyme's active site consists of five subsites for hyalobiuronate residues. In the terminology of Schechter, I., and Berger, A. ((1966) Biochemistry 5, 3371), these are s2-s1-s' 2-s3, where the reducing terminus is to the right, and cleavage occurs between s1 and s' 1. It is proposed that subsite s'2 has a high affinity for a substrate residue, while s1 and s'1 have low substrate affinity, and s2 and s' 3 are intermediate in affinity. This proposal is seen to have mechanistic implications. The reactions of several substrates show similar bell-shaped pH dependences, with optima in the region of pH 5 to 5.5. 相似文献
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Excess amino acid polymorphism in mitochondrial DNA: contrasts among genes from Drosophila, mice, and humans 总被引:13,自引:3,他引:10
Recent studies of mitochondrial DNA (mtDNA) variation in mammals and
Drosophila have shown an excess of amino acid variation within species
(replacement polymorphism) relative to the number of silent and replacement
differences fixed between species. To examine further this pattern of
nonneutral mtDNA evolution, we present sequence data for the ND3 and ND5
genes from 59 lines of Drosophila melanogaster and 29 lines of D. simulans.
Of interest are the frequency spectra of silent and replacement
polymorphisms, and potential variation among genes and taxa in the
departures from neutral expectations. The Drosophila ND3 and ND5 data show
no significant excess of replacement polymorphism using the
McDonald-Kreitman test. These data are in contrast to significant
departures from neutrality for the ND3 gene in mammals and other genes in
Drosophila mtDNA (cytochrome b and ATPase 6). Pooled across genes, however,
both Drosophila and human mtDNA show very significant excesses of amino
acid polymorphism. Silent polymorphisms at ND5 show a significantly higher
variance in frequency than replacement polymorphisms, and the latter show a
significant skew toward low frequencies (Tajima's D = -1.954). These
patterns are interpreted in light of the nearly neutral theory where mildly
deleterious amino acid haplotypes are observed as ephemeral variants within
species but do not contribute to divergence. The patterns of polymorphism
and divergence at charge-altering amino acid sites are presented for the
Drosophila ND5 gene to examine the evolution of functionally distinct
mutations. Excess charge-altering polymorphism is observed at the carboxyl
terminal and excess charge-altering divergence is detected at the amino
terminal. While the mildly deleterious model fits as a net effect in the
evolution of nonrecombining mitochondrial genomes, these data suggest that
opposing evolutionary pressures may act on different regions of
mitochondrial genes and genomes.
相似文献
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The cauliflower (Brassica oleracea L. var. botrytis) Or gene is a semi-dominant, single-locus mutation that induces the accumulation of high levels of beta-carotene in various tissues of the plant, turning them orange. As part of a map-based cloning strategy, molecular mapping of the Or gene in the cauliflower genome was undertaken in a mapping population consisting of 195 F2 individuals. By using amplified fragment length polymorphism (AFLP) in conjunction with bulked segregant analysis, we identified 10 AFLP markers closely linked to the Or gene. Four of the most closely linked flanking markers were converted into restriction fragment length polymorphism (RFLP) markers. Mapping of these markers in the mapping population placed two of them at 0.5 cM from the Or locus on one side, while another marker flanked the Or gene at 1.6 cM on the other side. Three of these markers were also successfully converted into sequence-characterized amplified region (SCAR) markers. These PCR-based markers will be useful for a large-scale application in facilitating the positional cloning of the Or gene. 相似文献
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The distribution of complex carbohydrates has been investigated at the light and electron microscope levels in sweat glands of normal subjects and patients with Hurler's or Hunter's disease. Normal sweat glands examined with a battery of light microscopic histochemical methods revealed sulphated complex carbohydrate in secretory granules of the dark cells. These granules lacked affinity for dialysed iron (DI) at the light and electron microscope levels. The DI method demonstrated acid complex carbohydrates ultrastructurally on the surface of the intercellular canaliculi and central lumen in normal sweat glands. DI-reactive acidic material, presumably of mucopolysaccharide nature, surrounded and extended between collagen bundles in the stroma of normal skin, but was absent from the band which ensheathed the sweat gland and consisted of individual rather than bundled collagen fibrils. DI-reactive mucopolysaccharide lined and partially filled vacuoles of dark cells showing a laminar distribution in vacuoles of clear cells in sweat glands of a Hunter patient. The DI method also visualized mucopolysaccharide distributed throughout vacuoles in fibroblasts of this patient. DI-reactive acid material covered the luminal surface of the sweat gland, coated collagen bundles in the stroma and spared the periglandular collagenous sheath in skin from Hurler and Hunter patients as in that from normal controls. Acid phosphatase was localized ultrastructually in vacuoles and nearby cytoplasm and on plasmalemmae of clear cells, dark cells and myoepithelial cells of sweat glands from Hurler and Hunter patients. Vacuoles of dermal fibroblasts and Schwann cells in these specimens also exhibited strong acid phosphatase activity. 相似文献
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Background and objectives The purpose of this paper is to describe the use of resident performance on an observed structured clinical examination (OSCE) as a tool to refine a mood disorders curriculum, and to disseminate a mood disorders OSCE for use in other residency settings.Methods A depression-focused OSCE and a direct observation evaluation tool were developed and implemented. A total of 24 first-year family medicine residents (PGY1) participated in the OSCE, and their performance was used to direct changes in a mood disorders curriculum.Results Residents performed well on general interview behaviours, and 67% were able to uncover depression in a patient presenting with headaches. Less than 50% of the residents asked about suicidal ideation and recreational drug use. Curriculum was added that addressed the latter deficiencies.Conclusions Tracking of resident performance on specific behaviours during OSCE sessions can be used for curriculum evaluation purposes. The mood disorders curriculum in additional family medicine residency programmes can now be evaluated using our depression-focused OSCE and Clinical Performance Checklist. 相似文献