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R C Siegel

Publications and source records attributed to R C Siegel.

15 recordsLinked to original sources
PubMed1979

Native cross-links in collagen fibrils induce resistance to human synovial collagenase.

A model system consisting of highly purified lysyl oxidase and reconstituted lathyritic chick bone collagen fibrils was used to study the effect of collagen cross-linking on collagen degradation by mammalian collagenase. The results indicate that synthesis of approx. 0.1 Schiff-base cross-link per collagen molecule results in a 2--3-fold resistance to human synovial collagenase when compared with un-cross-linked controls or samples incubated in the presence of beta-aminopropionitrile to inhibit cross-linking. These results confirm previous studies utilizing artificially cross-linked collagens, or collagens isolated as insoluble material after cross-linking in vivo, and suggest that increased resistance to collagenase may be one of the earliest effects of cross-linking in vivo. The extent of intermolecular cross-linking among collagen fibrils may provide a mechanism for regulating the rate of collagen catabolism relative to synthesis in normal and pathological conditions.

Aminopropionitrile↗
PubMed1979

Oxidative deamination of epsilon-aminolysine residues and formation of Schiff base cross-linkages in cell envelopes of Escherichia coli.

Oxidative deamination of the epsilon-amino group of lysyl residues to form allysine is the initial reaction in the cross-linking of collagen and elastin in vertebrates. The allysyl residues, generated by lysyl oxidase in this reaction, condense with either other allysyl residues or epsilon-amino groups of lysyl or hydroxylysyl to form aldol or Schiff base cross-links. This paper presents evidence that similar allysyl residues and Schiff base cross-links are synthesized in cell envelopes of Escherichia coli. Acid hydrolysis followed by amino acid analysis of envelopes either reduced with NaB[3H]4 or labeled with [14C]lysine and reduced with NaBH4 yielded allysine and two labeled fragments with elution profiles and molecular weights (250 and 330) consistent with Schiff base products derived at least in part from allysine. When [6-3H]lysine-labeled cell envelopes were incubated at 37 degrees C, gradual release of tritiated water occurred. This suggests that an enzymatic reaction catalyzes the deamination of lysine in E. coli membranes and that the higher molecular weight proteins detected in stationary phase or in log phase cell envelopes after NaBH4 reduction occur as a result of formation of Schiff base cross-links.

Amines↗
PubMed1979

Lysyl oxidase.

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Amino Acid Oxidoreductases↗
PubMed1979

Aortic aneurysm in Marfan's syndrome: changes in the ultrastructure and composition of collagen.

Aneurysmal aortic tissue and the mitral valve of a patient with Marfan's syndrome were examined. Biochemical analysis of the tissue showed a qualitative and quantitative defect in alpha 2 chain production of Type I collagen. On polyacrylamide gel electrophoresis of the aortic extract, two separate bands in the alpha 2 region and an increase of the alpha 1 to alpha 2 ratio were found. Examination by electron microscopy revealed elastic fibre degeneration, helical collagen fibres, and metabolically active modified smooth muscle cells. The formation of helical collagen fibres is attributed to a defect in the development of chains and cross-links of collagen precursors produced by the hypertrophic smooth muscle cells. Elastic fibre disintegration is believed to be due to a lack of support by Type I collagen fibres, which have decreased tensile strength. A scheme for the pathogenesis of aortic aneurysm and other connective tissue abnormalities in Marfan's syndrome is proposed as follows. Type I collagen fibres have decreased tensile strength because of a defect in the alpha 2 chain biosynthesis and decreased cross-linking. Over many years the wall of the ascending aorta is subjected to cyclic stresses and it dilates. Elastic fibres disintegrate. The attempt at repair by metabolically activated modified smooth muscle cells is abortive, and rupture is likely to occur.

Adult↗
PubMed1978

Biochemical and immunochemical study of lysyl oxidase in experimental hepatic fibrosis in the rat.

Lysyl oxidase catalyzes the crosslinking of collagen and elastin. Lysyl oxidase activity was measured and localized in rat liver during the evolution of hepatic fibrosis induced by CCl4. Enzyme activity measured with DL-[6-3H]-lysine-labeled collagen substrates in liver and plasma increased sharply after approximately 3 wk of injection, reached a maximum at 6 wk, and then decreased. The increase in activity correlated histologically with early connective tissue septa formation, and the magnitude of increase was significantly greater than that found for the intracellular collagen biosynthetic enzymes protocollagen prolyl hydroxylase and lysyl hydroxylase. Indirect immunofluorescence studies showed that lysyl oxidase was present in association with collagen in the extracellular space. However, it was not possible to correlate the distribution pattern with a particular liver cell type. These observations suggest that serial measurements of lysyl oxidase activity in liver or plasma may be useful for correlating changes in connective tissue formation with histologic connective tissue deposition.

Amino Acid Oxidoreductases↗
PubMed1977

Collagen cross-linking. Effect of D-penicillamine on cross-linking in vitro.

D-Pencillamine is believed to inhibit collagen cross-link biosynthesis by forming thiazolidine rings with lysyl-derived aldehydes that are intermediates in bifunctional cross-link synthesis. Recently, we showed that aldehyde biosynthesis catalyzed by lysyl oxidase occurs after the onset of fibril formation and that nascent aldehydes form Schiff-base cross-links rapidly in fibrils. This suggested that the accessibility of D-penicillamine to most aldehydes formed during cross-link synthesis might be limited. To study this, reconstituted chick bone collagen fibrils were incubated in vitro with highly purified lysyl oxidase and D-penicillamine. As reported in previous studies in vivo, allysine content increased and polyfunctional cross-link synthesis decreased with D-penicillamine. However, the concentration of bifunctional cross-links increased rather than decreased due to a 2-fold increase in N6:6'-dehydro-5,5'-dihydroxylysinonorleucine. Hydroxyallysine, an intermediate in formation of this Schiff base, decreased. A time study indicated that allysine levels increased primarily after the bulk of Schiff base synthesis. These results indicate that D-penicillamine does not inhibit bifunctional cross-link synthesis as previously suggested. Its principal effect is to block synthesis of polyfunctional cross-link products from Schiff base cross-link precursors and to cause accumulation of these precursors. This effect may be due to interference with the close molecular packing required for polyfunctional cross-link synthesis. These results also suggest a mechanism for the relative insensitivity of tissues such as bone with high hydroxylysine content to D-penicillamine. In this study, D-penicillamine caused selective accumulation of allysyl and not hydroxyallysyl residues. In bone as opposed to soft tissues, hydroxyallysyl residues are intermediates in synthesis of almost all cross-links.

Animals↗
PubMed1977

Scleroderma.

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Adult↗
PubMed1976

Collagen cross-linking. Purification and substrate specificity of lysyl oxidase.

Lysyl oxidase is a specific amine oxidase that catalyzes the formation of aldehyde cross-link intermediates in collagen and elastin. In this study, lysyl oxidase from embryonic chick cartilage was purified to constant specific activity and a single protein band on sodium dodecyl sulfate acrylamide gel electrophoresis. This band had an apparent molecular weight of 62,000. The eluted protein cross-reacted with inhibiting antisera developed against highly purified lysyl oxidase. The highly purified enzyme was active with both insoluble elastin and embryonic chick skin or bone collagen precipitated as reconstituted, native fibrils. There was low activity with nonhydroxylated collagen, collagen monomers, or native fibrils isolated from lathyritic calvaria. The maximum number of aldehyde intermediates formed per molecule of collagen that became insoluble was two. These results indicate that lysyl oxidase has maximum activity on ordered aggregates of collagen molecules that may be overlapping associations of only a few collagen molecules across. Formation of aldehyde intermediates and cross-links during fibril formation may facilitate the biosynthesis of stable collagen fibrils and contribute to increased fibril tensile strength in vivo.

Amino Acid Oxidoreductases↗
PubMed1976

Collagen cross-linking. Synthesis of collagen cross-links in vitro with highly purified lysyl oxidase.

In this paper, the synthesis of collagen cross-links in vitro was investigated in a defined system consisting of highly purified chick cartilage lysyl oxidase and chick bone collagen fibrils. Cross-link synthesis in vitro was quite similar to the biosynthesis of collagen cross-links in vivo. Enzyme-dependent synthesis of cross-link intermediates and cross-linked collagen derived from lathyritic collagen occurred. The concentration of the two principal reducible cross-links, N6:6'-dehydro-5,5'-dihydroxylysinonorleucine and N6:6'-dehydro-5-hydroxylysinonorleucine, increased to a peak value of approximately two cross-links per molecule and then decreased. Synthesis of histidinohydroxymerodesmosine and a second polyfunctional cross-link of unknown structure began after synthesis of bifunctional cross-links was largely completed and proceeded linearly afterwards. Inhibition of lysyl oxidase after the bulk of bifunctional cross-link synthesis had occurred did not alter the rate of decrease in reducible cross-link concentration but did inhibit further histidinohydroxymerodesmosine synthesis. These results indicate that lysyl oxidase and collagen fibrils are the only macromolecules required for cross-link biosynthesis in vivo. It is likely that the decrease in reducible cross-links observed during fibril maturation results from spontaneous reactions within the collagen fibril rather than additional enzymatic reactions.

Amino Acid Oxidoreductases↗