Histidine-rich glycoprotein binds to human IgG and C1q and inhibits the formation of insoluble immune complexes.
Purification of the complement component C1q from human serum using an established method resulted in the copurification of two 30 kDa proteins with an N-terminal sequence identical to human histidine-rich glycoprotein (HRG). Therefore, to explore the possibility that HRG can interact with C1q, we examined the ability of 81 kDa (native) and the 30 kDa proteins (presumably proteolytic N-terminal fragments of HRG) to bind to C1q, using both ELISA and optical biosensor techniques. Both forms of HRG were found to bind to the human complement component C1q and also to purified human and rabbit IgG by ELISA. Kinetic analyses of the HRG-C1q and HRG-IgG interactions using the IAsys biosensor indicate two distinct binding sites with affinities Kd1 0.78 x 10(-8) M and Kd2 3.73 x 10(-8) M for C1q, and one binding site with affinity Kd 8.5 x 10(-8) M for IgG. Moreover, the fact that both native and 30 kDa HRG bind to C1q and to IgG suggests that the IgG and C1q binding regions on HRG are located in the 30 kDa N-terminal region of the HRG molecule. The Fab region of IgG is likely to be involved in the HRG-IgG interaction since HRG also bound to F(ab')2 fragments with an affinity similar to that seen with the complete IgG molecule. Interestingly, the binding between HRG and IgG was significantly potentiated (Kd reduced from 85.0 to 18.9 nM) by the presence of physiological concentrations of Zn2+ (20 microM). Conversely, the presence of Zn2+ weakened the binding of HRG to C1q (Kd increased from 7.80 to 29.3 nM). Modulation of these interactions by other divalent metal cations was less effective with relative potencies being Zn2+ > Ni2+ > Cu2+. An examination of the effect of native and 30 kDa HRG on the formation of insoluble immune complexes (IIC) between ovalbumin and polyclonal rabbit anti-ovalbumin IgG revealed that physiological concentrations of HRG can markedly inhibit IIC formation in vitro. The results show that human HRG binds to C1q and to IgG in a Zn2+-modulated fashion, and that HRG can regulate the formation of IIC in vitro, thus indicating a new functional role for HRG in vivo.
Gorgani
N N
NN
Division of Immunology and Cell Biology, The John Curtin School of Medical Research, Australian National University, Canberra, ACT.
Parish
C R
CR
Easterbrook Smith
S B
SB
Altin
J G
JG
D000595:Amino Acid Sequence; D000936:Antigen-Antibody Complex / Q000378:metabolism*; D001769:Blood; D015922:Complement C1q / Q000378:metabolism*; D003300:Copper / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology / Q000378:metabolism*; D007700:Kinetics; D009532:Nickel / Q000494:pharmacology; D010047:Ovalbumin / Q000276:immunology; D010446:Peptide Fragments / Q000737:chemistry; D011506:Proteins / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D012995:Solubility; D015032:Zinc / Q000494:pharmacology; D000099053:Histidine-Rich Glycoprotein
D000936:Antigen-Antibody Complex; D007074:Immunoglobulin G; D010446:Peptide Fragments; D011506:Proteins; D000099053:Histidine-Rich Glycoprotein; D003300:Copper; D009532:Nickel; D015922:Complement C1q; D010047:Ovalbumin; D004492:Edetic Acid; D015032:Zinc