We have produced recombinant gamma-chain variant fibrinogens, gamma 308K, gamma 3081, and gamma 308A simultaneously with wild-type fibrinogen, gamma 308N, by genetic protein engineering using Chinese hamster ovary cells. Although all three variant fibrinogens are a result of a single amino acid substitution, the aberrant gamma-chains of gamma 308K and gamma 3081 fibrinogens migrated faster than gamma 308N. Furthermore, plasmic digestion profiles were examined in the presence of 5 mM ethylene glycol-bis(beta-aminoethyl ether)-N,N,N,'N'-tetraacetic acid (EGTA) or 1 mM CaCl2. In the presence of EGTA, the three variant fibrinogens were digested into D-1 and D-2 fragments slightly faster than wild type. In addition, the D-2 fragment derived from gamma 308K was further digested into D-3 by plasmin much faster than that from gamma 308N. These data suggest that cleavage of gamma 356Lys-gamma 357Ala bond by plasmin in gamma 308K, gamma 308I, and gamma 308A is slightly accelerated and the gamma 302Lys-gamma 303Phe bond is cleaved by plasmin rapidly in only the gamma 308K variant. Furthermore, the substitution of Lys for gamma 308Asn results in the generation of a new plasmin cleavage site between gamma 308Lys and gamma 309Gly in the presence of EGTA. In conclusion, a substitution at residue gamma 308Asn may cause a conformational change in the gamma-chain of fragment D affecting polymerization and plasmin cleavage.*