Detection and characterization of a protein isoaspartyl methyltransferase which becomes trapped in the extracellular space during blood vessel injury.
Injury to rat blood vessels in vivo was found to release intracellular pools of protein D-aspartyl/L-isoaspartyl carboxyl methyltransferase (PIMT) into the extracellular milieu, where it becomes trapped. This trapped cohort of PIMT is able to utilize radiolabeled S-adenosyl-L-methionine (AdoMet) introduced into the circulation to methylate blood vessel proteins containing altered aspartyl residues. As further shown in this study, methylated substrates are detected only at the specific site of injury. In vitro studies more fully characterized this endogenous PIMT activity in thoracic aorta and inferior vena cava. Methylation kinetics, immunoblotting, and the lability of methylated substrates at mild alkaline pH were used to demonstrate that both types of blood vessel contain an endogenous protein D-aspartyl/L-isoaspartyl carboxyl methyltransferase (PIMT). At least 50% of the PIMT activity is resistant to nonionic detergent extraction, suggesting that the enzyme activity becomes trapped within or behind the extracellular matrix (ECM). Quantities of lactate dehydrogenase (LDH), another soluble enzyme of presumed intracellular origin, were found to be similarly trapped in the extracellular space of blood vessels.
Weber
D J
DJ
Department of Biochemistry and Biophysics, Oregon State University, Corvallis 97331, USA.
McFadden
P N
PN
D000818:Animals; D001013:Aorta, Thoracic / Q000201:enzymology; D001808:Blood Vessels / Q000201:enzymology* / Q000293:injuries*; D003902:Detergents; D004730:Endothelium, Vascular / Q000166:cytology / Q000201:enzymology; D005110:Extracellular Space / Q000201:enzymology*; D005260:Female; D006863:Hydrogen-Ion Concentration; D007424:Intracellular Fluid / Q000201:enzymology; D007770:L-Lactate Dehydrogenase / Q000737:chemistry; D008745:Methylation; D009130:Muscle, Smooth / Q000166:cytology / Q000201:enzymology; D010047:Ovalbumin; D026601:Protein D-Aspartate-L-Isoaspartate Methyltransferase; D011489:Protein Denaturation; D011496:Protein Methyltransferases / Q000737:chemistry* / Q000378:metabolism / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D012436:S-Adenosylmethionine / Q000737:chemistry / Q000378:metabolism; D014020:Tissue Extracts / Q000737:chemistry; D014316:Tritium; D014682:Vena Cava, Inferior / Q000201:enzymology
D003902:Detergents; D014020:Tissue Extracts; D014316:Tritium; D012436:S-Adenosylmethionine; D010047:Ovalbumin; D007770:L-Lactate Dehydrogenase; D011496:Protein Methyltransferases; D026601:Protein D-Aspartate-L-Isoaspartate Methyltransferase