title: Carbamylation of erythrocyte membrane proteins: an in vitro and in vivo study.
abstract: OBJECTIVE
  To establish the degree of erythrocyte membrane protein carbamylation in uremic and nonuremic patients, and to characterize the in vitro binding of cyanate to the individual proteins of the cytoskeletal matrix.
  
  
  METHODS
  For in vivo studies, erythrocyte ghosts were digested with proteinase K and the released peptides colorimetrically assayed for carbamylation, using the diacetyl monoxime reagent, and quantitated using homocitrulline. For in vitro studies, erythrocyte ghosts were incubated with [14C] cyanate, and the membrane proteins separated by SDS-PAGE. Cyanate incorporation was quantitated by liquid scintillation counting and imaging densitometry of the excised bands.
  
  
  RESULTS
  Erythrocytes from uremic patients were found to have a greater level of carbamylation than those from nonuremic patients (47.09 +/- 7.80 and 25.89 +/- 6.92 nmol homocitrulline/mg proteolyzed protein released, respectively). In vitro incorporation of [14C] cyanate into membrane protein followed the sequence: spectrin > ankyrin > Band 4.1 > Band 3 > actin > Band 7.
  
  
  CONCLUSIONS
  The increased level of erythrocyte membrane protein carbamylation in uremic compared to nonuremic patients may lead to membrane destabilization and contribute to the decreased erythrocyte survival time observed in uremia.
authors.affiliation: Department of Chemistry and Biochemistry, University of Windsor, Ontario, Canada.
authors.forename: D J
authors.initials: DJ
authors.lastname: Trepanier
authors.forename: R J
authors.initials: RJ
authors.lastname: Thibert
authors.forename: T F
authors.initials: TF
authors.lastname: Draisey
authors.forename: P S
authors.initials: PS
authors.lastname: Caines
mesh_terms: D001345:Autoradiography; D002221:Carbamyl Phosphate / Q000378:metabolism*; D003485:Cyanates / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D019286:Endopeptidase K / Q000378:metabolism; D004910:Erythrocyte Membrane / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000378:metabolism*; D013054:Spectrophotometry, Atomic; D014511:Uremia / Q000378:metabolism
chemical_list: D003485:Cyanates; D008565:Membrane Proteins; D002221:Carbamyl Phosphate; D019286:Endopeptidase K