Isolation and sequence analyses of cDNA clones for the large subunits of two isozymes of rabbit calcium-dependent protease.
Two sets of cDNA clones were isolated from cDNA libraries prepared from poly(A+) RNA of rabbit lung and spleen by screening with the cDNA probe for the large subunit (80-kDa subunit) of chicken calcium-dependent protease (Ca2+-protease; Ohno, S., Emori, Y., Imajoh, S., Kawasaki, H., Kisaragi, M., and Suzuki, K. (1984) Nature 312, 566-570). The two sets of clones were identified as cDNA clones for two Ca2+-protease isozymes with high (mu-type) and low (m-type) calcium sensitivities from a comparison of the primary structures deduced from the nucleotide sequences with partial amino acid sequences from the two isozymes. The cDNA clones for the 80-kDa subunits of the mu- and m-type Ca2+-proteases contained, in total, about 1.5- and 2.2-kilobase cDNA inserts, respectively, which correspond roughly to the C-terminal halves of the coding regions and the entire 3'-noncoding regions. The two isozymes are encoded by two distinct mRNA species present in all the tissues examined, although the amount of mRNA significantly differs among the various tissues. Four E-F hand structures, typical calcium-binding structures in various calcium-binding proteins such as calmodulin, were detected in the C-terminal regions of both isozymes, as in the case of chicken Ca2+-protease. Comparison of the amino acid sequences of the two rabbit isozymes and the corresponding region of the chicken enzyme revealed marked homology, which indicates that these three enzymes have the same evolutionary origin. Furthermore, we suggest that the mu-type rabbit Ca2+-protease, rather than the m-type, is similar to chicken Ca2+-protease, which is regarded as an m-type enzyme in the C-terminal region. The evolution and molecular basis of the differences in calcium sensitivities of the Ca2+-proteases are discussed.
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002154:Calpain / Q000235:genetics*; D002645:Chickens; D004247:DNA / Q000235:genetics* / Q000302:isolation & purification; D004274:DNA, Recombinant; D007527:Isoenzymes / Q000235:genetics*; D008168:Lung / Q000032:analysis; D009693:Nucleic Acid Hybridization; D011061:Poly A / Q000235:genetics; D011487:Protein Conformation; D012313:RNA / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism; D011817:Rabbits; D013154:Spleen / Q000032:analysis
D004274:DNA, Recombinant; D007527:Isoenzymes; D012333:RNA, Messenger; D011061:Poly A; D012313:RNA; D004247:DNA; D002154:Calpain