The catalytically active serine protease domain of human complement factor I

Area of Science:

• Biochemistry
• Immunology
• Protease biochemistry

Background:

• Factor I (fI) is a key regulator of the complement system, acting as a protease.
• fI cleaves C3b and C4b, producing iC3b, a crucial opsonin, but requires cofactors like factor H (fH).
• The function of the fI heavy chain in catalysis remains unclear.

Purpose of the Study:

• To investigate the role of the fI heavy chain in complement regulation.
• To compare the catalytic activity of intact fI with its isolated SP domain.
• To elucidate the substrate specificity and cofactor dependence of fI-mediated cleavage.

Main Methods:

• Isolation of the intact SP domain of fI using partial proteolysis and affinity chromatography.
• Assay of amidolytic and proteolytic activities of fI and the SP domain.
• Investigation of cofactor (fH) dependence and inhibition by specific protease inhibitors (Pefabloc SC, antipain, suramin).

Main Results:

• Intact fI and the isolated SP domain exhibited similar amidolytic activities.
• fI cleaved C3(NH(3)) specifically at two sites in the presence of fH.
• The SP domain cleaved C3(NH(3)) at multiple sites without fH, and fH inhibited this activity.
• Suramin selectively inhibited fI but not the SP domain's proteolytic activity.

Conclusions:

• The fI heavy chain and cofactor fH are essential for orienting natural substrates and ensuring specific cleavage of C3b.
• These components restrict fI's catalytic activity to produce iC3b, highlighting a highly specific enzymatic mechanism.
• The SP domain alone possesses broader, less specific proteolytic activity compared to intact fI.