Abstract
As a result of blood vessel injury, protein D-aspartyl/L-isoaspartyl carboxyl methyltransferase (PIMT), a normally intracellular enzyme, becomes trapped within the meshwork of the vascular extracellular matrix where it can methylate substrate proteins. In this investigation we examined the distribution of such altered aspartyl-containing substrate proteins in the vascular wall. Nearly 90% of all the altered aspartyl residues were inaccessible to intracellular PIMT. Proteins of the extracellular matrix were found to be the major repository of altered aspartyl-containing polypeptides in the blood vessel wall, accounting for ∼70% of the total amount. Proteolytic cleavage of extracellular matrix proteins with cyanogen bromide (CNBr) revealed that collagens account for most of the altered aspartyl-containing proteins of the ECM. As a consequence of blood vessel injury, both type I and type III collagen along with other proteins were found to become methylated by injury-released PIMT. It is estimated that 1 cm of vein contains on the order of 5×1014 altered aspartyl residues involving between 1% and 5% of the total extracellular protein.
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REFERENCES
Adelstein, R. S., and Kuchl, M. W. (1970). Structural studies on rabbit skeletal actin. I. Isolation and characterization of the peptide produced by cyanogen bromide cleavage, Biochemistry 9, 1355–1364.
Aswad, D. W. (1995). Deamidation and Isoaspartate Formation in Peptides and Proteins, CRC Press, Boca Raton, Florida.
Bhown, A. S., Mole, J. E., et al. (1978). Methanol solvent system for rapid analysis of phenylthiohydantoin amino acids by high-pressure liquid chromatography, J. Chromatog. 148, 532–535.
Bornstein, P., and Sage, H. (1980). Structurally distinct collagen types, Annu. Rev. Biochem. 49, 957–1003.
Brennan, T. V., and Clarke, S. (1995). Deamidation and isoaspartate formation in model synthetic peptides: The effects of sequence and solution environment, in Deamidation and Isoaspartate Formation in Peptides and Proteins (Aswad, D. W., ed.), CRC Press, Boca Raton, Florida, pp. 66–88.
Chazin, W. J., and Kossiakoff, A. A. (1995). The role of secondary and tertiary structures in intramolecular deamidation of proteins, in Deamidation and Isoaspartate formation in Peptides and Proteins (Aswad, D. W., eds.), CRC Press, Bocca Raton, Florida, pp. 194–206.
DiAugustine, R. P., Gibson, B. W., et al. (1987). Evidence for isoaspartyl forms of mouse epidermal growth factor, Anal. Biochem. 165, 420–429.
Friedman, A. R., Ichhpurani, A. K., et al. (1991). Degradation of growth hormone releasing factor analogs in neutral aqueous solution is related to deamidation of asparagine residues, Int. J. Peptide Protein Res. 37, 14–20.
George-Nascimento, C., Gyenes, A., et al. (1988). Characterization of recombinant human growth factor produced in yeast, Biochemistry 27, 797–802.
Gilbert, J. M., Fowler, A., et al. (1988). Purification of homologous protein carboxyl methyltransferase isozymes from human and bovine erythrocytes, Biochemistry 27, 5227–5233.
Green, G. D., and Reagan, K. (1992). Determination of hydroxyproline by high performance liquid chromatography, Anal. Biochem. 201, 265–269.
Harris, J. U., Robinson, D., and Johnson, A. J. (1980). Analysis of (3-phenyl,2-thio)hydantoin amino acids by high-performance liquid chromatography, Anal. Biochem. 105, 239–245.
Humphries, M. J. (1993). Peptide sequences in matrix proteins recognized by adhesion receptors, in Cell Biology: A Series of Monographs, Academic Press, San Diego, pp. 289–308.
Jaffe, E. A., Nachman, R. L., et al. (1973). Synthesis of antihemophilic factor antigen by cultured human endothelial cells, J. Clin. Invest. 52, 2757.
Johnson, B. A., and Aswad, D. W. (1990). Identities, origins and metabolic fates of the substrates for eukaryatic protein carboxyl methyltransferases, in Protein Methylation (Paik, W. S., and Kim, S. eds.), CRC Press, Boca Raton, Florida, pp. 196–207.
Johnson, B. A., Langmack, E. L., and Aswad, D. W. (1987). Partial repair of deamidation-damaged calmodulin by protein carboxyl methyltransferase, J. Biol. Chem. 262, 12283–12287.
Johnson, D. J., LaBourene, J., et al. (1993). Relative efficiency of incorporation of newly synthesized elastin and collagen into aorta, pulmonary artery and pulmonary vein of growing pigs, Connect. Tiss. Res. 29, 213–221.
Maurel, E., Azema, C., et al. (1990). Collagen of the normal and the varicose human saphenous vein: A biochemical study, Clin. Chim. Acta 193, 27–38.
Morton, L. F., and Barnes, M. J. (1982). Collagen polymorphism in the normal and diseased blood vessel wall, Atherosclerosis 42, 41–51.
Murata, K., and Motoyama, T. (1990). Collagen species in various sized human arteries and their changes with intimal proliferation, Artery 17, 96–106.
Murata, K., Motayama, T., and Kokake, C. (1986). Collagen types in various layers of the human aorta and their changes with atherosclerotic process, Athereosclerosis 60, 251–262.
Ozols, J. (1991). Multiple forms of liver microsomal flavin-containing monooxygenases: complete covalent structure of form 2, Arch. Biochem. Biophys. 290, 103–15.
Sage, H., Pritzl, P., and Bornstein, P. (1980). A unique pepsin-sensitive collagen synthesized by aortic endothelial cells in culture, Biochemistry 19, 5747–5755.
Service, R. F. (1995). Designer tissues take hold, Science 270, 230–232.
Starcher, B. C., and Galione, M. J. (1976). Purification and comparison of elastins from different animal species, Anal. Biochem. 74, 441–447.
Sun, A. Q., Yuksel, K. U., and Gracy, R. W. (1992). Relationship between the catalytic center and primary degradation site of triosephosphate isomerase: Effects of active site modification and deamidation, Arch. Biochem. Biophys. 293, 382–390.
Takasago, T., Nkamua, K., et al. (1992). Analysis of collagen type III by uninterrupted sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting: Changes in collagentype III polymorphism in aging rats, Electrophoresis 13, 373–378.
Tanno, T., Yoshinaga, K., and Sato, T. (1993). Alteration of elastin in aorta of diabetics, Atherosclerosis 101, 129–134.
Venkatesh, Y. P., and Vithayathil, P. J. (1984). Isolation and characterization of monodeamidated derivatives of bovine pancreatic ribonuclease A, Int. J. Peptide Protein Res. 23, 494.
Weber, D. J., and McFadden, P. N. (1996a). Identification of proteins modified by protein (D-aspartyl/L-isoaspartyl) carboxyl methyltransferase, in Protein Protocols Handbook (Walker, J. M., ed.), Humana Press, Totawa, New Jersey, pp. 507–515.
Weber, D. J., and McFadden, P. N. (1996b). Protein methylation in the nervous system, in Neuromethods (Hemmings, H. C., ed.) Humana Press, Totawa, New Jersey
Weber, D. J., and McFadden, P. N. (1997). Detection and characterization of a protein isoaspartyl methyltransferase which becomes trapped in the extracellular space during blood vessel injury, J. Protein Chem., this issue.
Yamada, K. M. (1991). Fibronectin and other cell interactive glycoproteins, in Cell Biology of Extracellular Matrix, Plenum Press, New York.
Yurchenco, P. D., and O'Rear, J. (1993). Supramolecular organization of basement membranes, in Cell Biology: A Series of Monographs, Academic Press, San Diego, pp. 19–47.
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Weber, D.J., McFadden, P.N. Injury-Induced Enzymatic Methylation of Aging Collagen in the Extracellular Matrix of Blood Vessels. J Protein Chem 16, 269–281 (1997). https://doi.org/10.1023/A:1026352908978
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DOI: https://doi.org/10.1023/A:1026352908978