Fibrinogen assembly and secretion. Role of intrachain disulfide loops.
Human fibrinogen is a homodimer composed of three different (Aalpha, Bbeta, gamma) polypeptide chains. The chains are linked by 29 inter- and intrachain disulfide bonds. Each half-molecule has 6 intrachain disulfide bonds, which form loops in the carboxyl-terminal region of each of the chains. Aalpha chain has one disufide loop (Cys442-Cys472), Bbeta has three (Cys201-Cys286, Cys211-Cys240, and Cys394-Cys407), and gamma has two loops (Cys153-Cys182 and Cys326-Cys339). The intrachain loops are conserved in fibrinogens of different species. We changed, by site-directed mutagenesis, the cysteines, which form the intrachain loops, to serine or alanine. Fibrinogen chain assembly and secretion was determined in transiently transfected COS cells expressing two normal and a mutant fibrinogen chain. In the Bbeta and gamma chains, disruption of the disulfide loops closest to the "coiled-coil" region (CysBbeta211-Cys240, CysBbeta201-Cys286, and Cysgamma153-Cys182) abolished chain assembly and secretion, indicating that the disulfide loops closest to the coiled-coil region are essential for chain assembly. By contrast, preventing formation of the disulfide loops, which are toward the carboxyl termini of each of the chains, had different effects. Disruption of the single Aalpha disulfide loop had no effect, as did disruption of BbetaCys394-Cys407. However, disruption of Cysgamma326-Cys339, which is similar in size and location to CysBbeta394-Cys407, allowed chain assembly to occur, but the assembled chains were not secreted.
Zhang
J Z
JZ
Lindsley F. Kimball Research Institute, New York Blood Center, New York, New York 10021, USA.
Redman
C
C
D000818:Animals; D019556:COS Cells; D004220:Disulfides / Q000737:chemistry*; D005340:Fibrinogen / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D016297:Mutagenesis, Site-Directed; D011499:Protein Processing, Post-Translational
D004220:Disulfides; D005340:Fibrinogen