J. Biol. Chem., Vol. 258, Issue 6, 3446-3452, Mar, 1983
Secretion of proalbumin by canavanine-treated Hep-G2 cells
CM Redman, G Avellino and S Yu
The two processing sites in the conversion of preproalbumin to albumin are
marked by arginine residues. Therefore, to study the mechanisms of albumin
processing and secretion, the arginine residues of nascent albumin were
replaced with canavanine by the incubation of Hep-G2 cells with this
arginine analog. During a 4-h interval, canavanine inhibited (67%) the
secretion of nascent albumin and increased the intracellular transit time
of albumin secretion from 24 to 39 min. At 1 h, canavanine inhibited total
protein synthesis by 19% and albumin synthesis by about 40%. Both the
intracellular and secreted albumin produced by canavanine- treated cells
were analyzed by DEAE-cellulose chromatography and were found to be more
acidic than normal proalbumin and albumin. Further analysis on sodium
dodecyl sulfate polyacrylamide gel electrophoresis indicated that the
albumin produced and secreted by canavanine-treated cells appeared to have
a larger molecular weight (by 4000) than serum albumin. The
canavanine-treated cells were incubated with L-[3H]leucine and
L-[3H]phenylalanine and the location of radioactive L-leucine and L-
phenylalanine in the 30 NH2-terminal amino acid residues of secreted
albumin was determined. The results indicated that canavanine-treated cells
secreted proalbumin (79%) and also some fully processed albumin (21%).
Preproalbumin was not secreted. Untreated Hep-G2 cells mostly secreted
fully processed serum albumin (93%) with only traces of proalbumin (7%).
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