Cale. Tiss. Res. 4 (Supplement), 42 (1970)

Enzymatic Cross-linking of Bone Collagen

1%. C. SIEGEL and G. R. 5IA~mI~

National Institute of Dental Research, NItt , Bethesda, Md

Exper imenta l la thyrism has received extensive s tudy since it is an acqui- red disease tha t may be an experimental model of congenital or acquired human connective tissue disease processes. Animals fed lathyrogens such as /~-aminopropionitrile (BAPST) develop connective tissue disorders such as dissecting aortic aneurysms, scolioses and exostoses. These lesions arise from weak connective tissue as a result of the inhibition of collagen and elastin cross-linking. In cross-linking specific lysyl residues in collagen and elastin are converted to peptide bound a-aminoadipic-~-semialdehyde (ally- sine) by deaminat ion and oxidation of the s-carbon a tom in lysine. In col- lagen, these lysine derived aldehydes can then react to form cross-links by either an aldol condensation with a second allysyl residue or by Schiff base format ion with the s-amino group of other lysyl or hydroxylysy l residues. We have recently demonstra ted tha t the format ion of allysine in bone col- lagen is catalyzed by an enzyme, lysyl oxidase. The present work describes the properties of lysyl oxidase and its action on the collagen molecule in vitro.

When part ial ly purified lysyl oxidase is incubated with la thyri t ic chick bone collagen, enzyme dependent cross-linking of the collagen is observed. There is formation of the /~1~ collagen component, the cross-linked dimer of an a l and a2 chain, and product ion of insoluble collagen. Allysine is pro- duced in both the a l and a2 chains from the lysyl residue closest to the amino terminal end in both cases. Ei ther a lysyl or an hydroxylysy l residue in this position can be converted to the corresponding aldehyde by lysyl oxidase. This region of the collagen molecule is unusual since it does not have the typical helical s tructure of collagen.

We have detected lysyl oxidase act iv i ty in extracts of chick, guinea pig, mouse and fetal calf bone. The enzyme is irreversibly inhibited b y physiologically active levels of BAPST. ~ o enzyme act iv i ty is detectable in bone extracts prepared from copper deficient chicks. Proteins other than collagen and elastin have not been found to be substrates for lysyl oxidase. Since lysyl oxidase is specific for those lysyl residues in collagen tha t are conver ted to allysine in vivo and it is inhibited by low levels of BAPbT, we conclude tha t lysyl oxidase is the enzyme active in cross-linking in vivo. In lathyrism, lysyl oxid~se act ivi ty is impaired. As a result~ there is insuf- ficient allysine formed for normal cross-linking and therefore, the observed connective tissue lesions occur.