Isolation and some physicochemical properties of requiem shark myosin.

cDNA libraries were constructed from fast skeletal muscles of carp acclimated to 10 and 30 degrees C for a minimum of 5 weeks and were screened for myosin alkali light chains, LC1 and LC3, using an anti-skipjack LC1 polyclonal antibody. Two types of LC1 cDNA clone were isolated and termed LC1a and LC1b: their nucleotide sequences showed 92% homology. The ratio of LC1a to LC1b cDNA clones isolated was approximately 3:1, showing no apparent changes following temperature acclimation. The occurrence of the two isoforms was further confirmed by N-terminal amino acid sequencing of purified LC1. No isoform was, however, detected for LC3, while homology in the overlapping region between LC1a and LC3 cDNAs was only 65% even after the most probable alignment. Southern blot analyses probed with cDNA clones specific to LC1a and LC3 showed different hybridization patterns from each other, demonstrating that carp LC1 and LC3 are encoded by different genes. These results are in marked contrast to those from higher vertebrates which express LC1 and LC3 from a single gene by alternative RNA transcription and two modes of splicing. Northern blot analysis showed that the ratios of LC3/LC1 mRNAs were significantly higher (3.93) in 30 degrees C-acclimated than in 10 degrees C-acclimated (3.10) carp.

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Monoclonal Antibodies Raised against Carp Myosin

Nakaya

Watabe

1994

Fisheries science

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Several cell lines producing monoclonal antibodies against carp fast muscle myosin were obtained by fusion of immunized BALB/c mouse spleen cells with syngeenetic NS-1 myeloma cells and subsequent cloning by the limiting dilution method. Monoclonal antibodies, which were obtained when myosin was injected to mice as an antigen, recognized myosin subfragment-1 (S1). Further assignments for antigenic sites in the S1 molecule revealed that antibodies from 35A11 and 41C2 cell lines were specific to 50 and 25 kDa fragments of S 1 tryptic digests, respectively. No cell line producing monoclonal antibody specific to the 20 kDa fragment of S 1 tryptic digests was established. On the other hand, cell lines for monoclonal antibodies against myosin rod were established only when the rod was administered to mice as an antigen. The 37G3 cell line produced the antibody specific to myosin rod and reacted with its light meromyosin part. However, all monoclonal antibodies thus raised failed to discriminate between two types of myosin from carp acclimated to 10 and 30•Ž, irrespective of myosin S1 or rod.

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Isolation and some physicochemical properties of requiem shark myosin.

Cited by 5 publications

References 1 publication

Effects of urea on actin-activated Mg2+-ATPase of requiem shark myosin

Effects of urea on actin-activated Mg2+-ATPase of requiem shark myosin

The Two Essential Light Chains of Carp Fast Skeletal Myosin, Lc1 and Lc3, are Encoded by Distinct Genes and Change their Molar Ratio Following Temperature Acclimation

Monoclonal Antibodies Raised against Carp Myosin

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