Functional effects of domain deletions in a multidomain serine protease, C1r.
The C1r subcomponent of the first component of complement is a complex, multidomain glycoprotein containing five regulatory or binding modules in addition to the serine protease domain. To reveal the functional role of the N-terminal regulatory domains, two deletion mutants of C1r were constructed. One mutant comprises the N-terminal half of domain I joined to the second half of the highly homologous domain III, resulting in one chimeric domain in the N-terminal region, instead of domains I-III. In the second mutant most of the N-terminal portion of domain I was deleted. Both deletion mutants were expressed in the baculovirus-insect cell expression system with yields typical of wild type C1r. Both mutants maintained the ability of the wild type C1r to dimerize. The folding and secretion of the recombinant proteins was not affected by these deletions, and C1-inhibitor binding was not impaired. The stability of the zymogen was significantly decreased however, indicating that the N-terminal region of the C1r molecule contains essential elements involved in the control of activation of the serine protease module. Tetramer formation with C1s in the presence of Ca2+ was abolished by both deletions. We suggest that the first domain of C1r is essential for tetramer formation, since the deletion of domain I from C1r impairs this interaction.
Cseh
S
S
Institute of Enzymology, Biological Research Center, Hungarian Academy of Sciences, Budapest, Hungary.
Gál
P
P
Sárvári
M
M
Dobó
J
J
Lörincz
Z
Z
Schumaker
V N
VN
Závodszky
P
P
D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D003174:Complement C1 Inactivator Proteins / Q000378:metabolism; D015923:Complement C1r / Q000737:chemistry / Q000502:physiology*; D018076:DNA, Complementary / Q000302:isolation & purification; D008969:Molecular Sequence Data; D009154:Mutation; D011994:Recombinant Proteins / Q000096:biosynthesis; D018411:Spodoptera; D013329:Structure-Activity Relationship
D003174:Complement C1 Inactivator Proteins; D018076:DNA, Complementary; D011994:Recombinant Proteins; D015923:Complement C1r; D002118:Calcium