Rat major acute-phase protein: biosynthesis and characterization of cDNA clone.
The major acute-phase protein (alpha 1-MAP) of rat serum is induced in response to inflammation. This induction may be attributed to a corresponding increase in the level of translatable mRNA for the protein. Using in vitro and in vivo systems, various biosynthetic processing intermediates of this glycoprotein have been isolated. alpha 1-MAP is translated in a rabbit reticulocyte system as a preprotein with an amino-terminal signal peptide and an apparent molecular weight of 51,000. Translation of rough microsomes yields a product with a mass of 57,000 Da, representing the core glycosylated form of alpha 1-MAP. Cotranslational glycosylation appears to occur in a stepwise fashion, since three glycosylated forms of alpha 1-MAP (51,000, 54,000, and 57,000 Da) were detected in polysome translations; these products were digested by endoglycosidase H to a 48,000-Da protein. Two intracellular forms of alpha 1-MAP were observed in vivo, a 57,000-Da (core carbohydrate sidechains) and a 66,000-Da protein (mature complex carbohydrate side-chains); the latter was the only component secreted into the culture medium. To extend our studies on this protein, a cDNA clone specific for alpha 1-MAP was isolated. The recombinant was positively identified by hybrid selection procedures and contains a 1.55-kb insert. Partial radiosequence analysis of the primary translation product indicated the distribution of Leu, Ile, Cys, and Met in the amino-terminal region of this protein. To relate the location of these amino acids with the nucleotide sequence, cDNA was analyzed by the method of Maxam and Gilbert. These results indicate that the cDNA insert contains the 3' poly(A) tail, and alignment of the 5' end of the cDNA with the available amino acid sequence of the primary translation product corroborated that the insert encodes the entire alpha 1-MAP protein except for the first four amino acids of the signal peptide.
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002474:Cell-Free System; D003001:Cloning, Molecular; D004247:DNA / Q000302:isolation & purification*; D008862:Microsomes, Liver / Q000378:metabolism; D008970:Molecular Weight; D011132:Polyribosomes / Q000378:metabolism; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D012333:RNA, Messenger / Q000302:isolation & purification; D051381:Rats; D011919:Rats, Inbred Strains; D000511:alpha-Macroglobulins / Q000096:biosynthesis* / Q000235:genetics
D012333:RNA, Messenger; D000511:alpha-Macroglobulins; D004247:DNA