title: The primary sequence and the subunit structure of mouse alpha-2-macroglobulin, deduced from protein sequencing of the isolated subunits and from molecular cloning of the cDNA.
abstract: Mouse plasma alpha-2-macroglobulin (m alpha 2M) was isolated and the N-terminal amino-acid sequences determined after separation of the 165-kDa and 35-kDa subunits. These sequences were compared to the protein sequence predicted by the cDNA, which was cloned from a mouse liver library and sequenced. From these data it is evident that both subunits are encoded by one mRNA of approximately 5 kb expressed predominantly in liver. The smaller subunit, with the N-terminal sequence DLSSSDLT, comprises the C-terminal 257 residues of m alpha 2M and is derived from a single-chain precursor probably by proteolytic processing at an arginine residue in the sequence PTRDLSS. Analysis of the predicted protein further showed all the salient features of a proteinase inhibitor of the macroglobulin family: a bait region that deviates from all known sequences in this family, a very conserved internal thiolester site and conserved cysteine residues and putative N-glycosylation sites. The synthesis of m alpha 2M in adult liver was demonstrated by Northern blotting and in fetal liver by in-situ hybridization. Transient transfection of COS cells with the cDNA under control of a viral promoter demonstrated the secretion and partial processing of m alpha 2M in the culture medium. In plasma the level of m alpha 2M was found to be stable as expected for the murine counterpart of human plasma alpha-2-macroglobulin. The possibilities of using the mouse as a genetic model to study this proteinase inhibitor in vivo are discussed.
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D003001:Cloning, Molecular; D004247:DNA; D004591:Electrophoresis, Polyacrylamide Gel; D017403:In Situ Hybridization; D051379:Mice; D008969:Molecular Sequence Data; D010957:Plasmids; D012313:RNA / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D000511:alpha-Macroglobulins / Q000235:genetics*
chemical_list: D000511:alpha-Macroglobulins; D012313:RNA; D004247:DNA