Modulation of clearance of recombinant serum albumin by either glycosylation or truncation.
Albumin is an abundant non-glycosylated plasma protein with a slow clearance profile. It has been employed as a fusion partner in efforts to slow the clearance of small antithrombotic proteins like hirudin. In the present study, the in vivo clearance of recombinant rabbit serum albumin (rRSA), of mutant rRSAs containing consensus sequences for N-linked glycosylation (D494N and V14T variants), and of mutant mini-proteins truncated at albumin domain boundaries (rRSAs 1-185, 1-377, or 378-584) was examined. Mean terminal catabolic half-lives (t(0.5)cat) in rabbits for plasma-derived RSA, rRSA, and the V14T variant did not differ significantly (range 4. 32-4.76 days). In contrast, mean t(0.5)cat was reduced to 2.87 days for the D494N variant and to less than 0.071 days for all mini-proteins. The mini-proteins were found in the urine in tissue distribution experiments, suggesting a renal route of clearance. Our results suggest that all three internally repeated albumin domains are required to maintain the slow in vivo clearance profile of albumin, and that albumin glycosylation can be associated with an acceleration of clearance. This information could be used to design fusion proteins, including those with antithrombotic properties, with predictably altered in vivo half-lives less than that of serum albumin.
Sheffield
W P
WP
Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada. sheffiel@mcmaster.ca
Marques
J A
JA
Bhakta
V
V
Smith
I J
IJ
D000818:Animals; D001345:Autoradiography; D004591:Electrophoresis, Polyacrylamide Gel; D006031:Glycosylation; D006207:Half-Life; D007267:Injections; D007457:Iodine Radioisotopes; D008297:Male; D016297:Mutagenesis, Site-Directed; D015202:Protein Engineering; D011817:Rabbits; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000493:pharmacokinetics; D012709:Serum Albumin / Q000737:chemistry / Q000235:genetics / Q000493:pharmacokinetics*; D013997:Time Factors; D014018:Tissue Distribution
D007457:Iodine Radioisotopes; D011994:Recombinant Proteins; D012709:Serum Albumin