Partial sequence of human complement component factor B: novel type of serine protease.

Abstract

Factor B (a component of the alternative pathway of complement) is believed to contain the proteolytic site of the complex enzymes C3 convertase (C3bB) and C5 convertase (C3bnB). Conflicting results have been obtained in regard to the inactivation of these enzymes by diisopropyl phosphorofluoridate but it has been suggested that activated Factor B (Factor B) is a serine protease with the active site in Bb, a COOH-terminal fragment of approximately 60,000 molecular weight. Partial amino acid sequence studies of Bb derived from human Factor B have shown that the NH2-terminal 40 residues have no homology with NH-terminal sequences of other serine proteases. However, positioning of a further 170 residues out of approximately 290 residues in two continuous CNBr fragments from the COOH terminus has shown that there is a strong homology of sequence in this section. The active site residues histidine, aspartic acid, and serine all are present in positions corresponding with those of typical serine proteases. It is suggested that Factor B is a novel type of serine protease with a catalytic chain of molecular weight twice that of proteases previously studied and probably with a different activation mechanism.In the alternative pathway of activation of complement, activated Factor B (Factor B) in association with C3b forms the complex proteases C3 convertase C3bB and C5 convertase (C3b)nB (1, 2). On activation by Factor D, Factor B, a glycoprotein of about 90,000 molecular weight, is split into an NH2-terminal fragment . Ba is about 30,000 molecular weight and Bb is about 60,000 molecular weight. Medicus et al. (6) reported that C3 and C5 convertases of both pathways of activation are inhibited by diisopropyl phosphorofluoridate (iPr2P-F) and that, if a radioactive reagent is used, it is found in the Bb fragment, suggesting that Factor B is a serine protease with the active center in Bb. An earlier report, however, showed the alternative pathway CS convertase to be resistant to inhibition by iPr2P-F (7). Because the suggested catalytic peptide Bb is also about twice the length of the catalytic peptides of all other serine proteases studied so far, it appeared that Factor B could not be a typical serine protease. The amino acid sequences of many of these proteases are known and they show strong homology with each other with many highly conserved features, notably in the NH2 terminus of the catalytic peptide and in the active site residues, suggesting that all have similar catalytic and activation mechanisms (8).Sequence studies have therefore been made of Bb and have shown that, although the essential features of the active site residues of serine proteases are present and are in similar positions relative to the COOH terminus, the NH2-terminal section, which is -u300 residues longer, does not contain the characteristic NH2-terminal sequence of other serine proteases. METHODSPurification of Factor B and Fragments Ba and Bb. Minor modifications were made to the procedure of Kerr (4) for the isolation of Factor B fr...

Human complement factor B: Functional properties of a recombinant zymogen of the alternative activation pathway convertase

“…For these sections, the pFB2 cDNA adds a novel coding sequence which taken together comprises 16 nucleotide residues. The peptides encoded by this added sequence are consistent with the protein sequencing data (15)(16)(17)(18).…”

“…The entire amino acid sequence of human factor B has been established combining both peptide sequencing and translation of the partial nucleotide sequences of cDNA and cosmid clones (14)(15)(16)(17)(18)(19). In this contribution, we present the first full-length cDNA transcript for human factor B and the functional analysis of the recombinant zymogen obtained by expressing this cDNA in COS cells.…”

Molecular Basis for the Microheterogeneity of Human Complement Factor B

“…cated in the C-terminal half of the Bb frag ment [2], The complete primary structure of factor B [3] allowed the precise location of the unique sulfhydryl group [4,5], the active-site His, Asp, and Ser residues [6], the Arg-Lys activation site [2,7], and four asparaginelinked glycosylation sites, two of each in the Ba and in the Bb fragments [2,5]. All carbo hydrates are N-linked, since the absence of galactosamine rules out the eventuality of Oglycosidic linkages [5].…”

Molecular Biology of the Human and Mouse MHC Class III Genes: Phylogenetic Conservation, Genetics and Regulation of Expression

“…Partial amino acid sequences had been published for factor B (Christie et al 1980), C4 (reviewed in Porter & Reid 1979) and C2 (Kerr & Gagnon 1982). In order to identify cDNA clones carrying factor B, C4, and C2 coding sequences, portions of their known protein sequences whose amino acid residues displayed a minimal complexity of codon usage were chosen.…”