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A second isoform of chicken brush border myosin I contains a 29‐residue inserted sequence that binds calmodulin
Author(s) -
Halsall David J.,
Hammer John A.
Publication year - 1990
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(90)80305-3
Subject(s) - calmodulin , complementary dna , microbiology and biotechnology , gene isoform , insert (composites) , biology , biochemistry , peptide sequence , alternative splicing , chemistry , enzyme , gene , mechanical engineering , engineering
Chicken brush border myosin I (CBB‐MI) is a single‐headed, nonfilamentous, myosin‐like mechanoenzyme which, as isolated, has 3 mol of calmodulin (CAM) ‘light chains’ bound per mole of 119 kDa heavy chain. We have isolated a partial cDNA clone for CBB‐MI that encodes the C‐terminal ~ 35 kDa of the heavy chain. The sequence of this clone is identical to that of an authentic, near‐full‐length CBB‐MI cDNA clone reported recently, except for an 87‐bp/29‐residue insertion occurring ~32 kDa from the C‐terminus. This insert, which is probably generated by an alternate splicing event, is expressed in brush border as part of a message of the size predicted for the CBB‐MI heavy chain, although the steady state level of this transcript is ~8‐fold lower than for transcripts lacking the insert. 125 I‐CAM overlays of this cDNA clone (expressed as a trpE fusion protein in E. coli ) indicate that it binds one more calmodulin than does a second cDNA clone that lacks the 29‐residue insert. A synthetic peptide corresponding to the insert sequence binds tightly to CAM‐Sepharose, demonstrates a shift and enhancement of fluorescence in the presence of CAM, and binds CAM in solution with a K D of 190 nM (in 100 mM KCl). We conclude that a second, low‐abundance isoform of CBB‐MI contains an additional (and possibly fourth) CAM binding site as a result of a 29‐residue peptide that is inserted into the tail domain by an apparent alternate splicing event.