Background: Cyanate formed spontaneously from urea carbamoylates non-protonated amino groups of protein, irreversibly altering function, charge and structure. Carbamoylated proteins in renal tissue have not been examined hitherto. Objectives: To identify homocitrulline (ε-amino-carbamoyl-lysine), a result of in vivo carbamoylation by urea-derived cyanate, from patients with renal disease or in newly transplanted kidneys by immunohistochemistry. To evaluate enzymatic activity of carbamoylated and non-carbamoylated matrix metalloproteinase-2 and correlate this with renal tissue carbamoylated in vivo. Design: Anti-homocitrulline antibody is specific for homocitrulline and was used to identify carbamoylation of ε-amino-lysine in renal biopsies from patients with elevated BUN, with isolated proteinuria, and as controls, from normal donors at time of transplantation. Enzymatic activity of matrix metalloproteinase-2 carbamoylated in vitro was evaluated. Results: Homocitrulline was present in glomerular basement membrane (8/10), mesangium (8/10), tubular epithelium and cytoplasm (7/10) and Bowman's capsule (1/10) in patients with elevated BUN. The discordant patterns of glomerular and tubular localization of homocitrulline versus immune complexes indicated that the carbamoylated proteins were not a component of immune deposits but were modified proteins in renal tissue. No homocitrulline was found in transplanted kidneys (14/15) or in proteinuric patients (2/2). Enzymatic activity of both human and rat matrix metalloproteinase-2 was strongly inhibited in a dose-dependent fashion when incubated with cyanate. Conclusions: In situ carbamoylation in proteins occurred in kidneys of patients with renal dysfunction but not in normal newly transplanted kidneys. Decreased enzymatic activity of carbamoylated enzymes may alter specific renal regulatory mechanisms. Carbamoylated proteins with altered function and charge may represent a previously underestimated mechanism in renal pathophysiology.
- Matrix metalloproteinase-2
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