Expression and biological activity of Baculovirus generated wild-type human slow α tropomyosin and the Met9Arg mutant responsible for a dominant form of nemaline myopathy

P. Anthony Akkari, Yuhua Song, Sarah Hitchcock-DeGregori, Lori Blechynden, Nigel Laing

Research output: Contribution to journalArticlepeer-review

26 Scopus citations

Abstract

We have previously reported a Met9Arg mutation in the human skeletal muscle α tropomyosin gene (TPM3) associated with autosomal dominant nemaline myopathy [Nat. Genet. 9 (1995) 75]. We describe here the generation of wild-type (Wt-tpm3) and Met9Arg (M9R-tpm3) mutant human skeletal muscle slow α tropomyosin using the Baculovirus expression vector system (BEVS). This system produces correct posttranslationally modified recombinant tropomyosin proteins in insect cells. We show that the interactions of Wt-tpm3 with actin and tropomyosin are comparable to those of fast α tropomyosin isolated from chicken striated muscle. However, the recombinant M9R-tpm3 is at least 100 times less effective at binding actin than Wt-tpm3. This paper represents the first study of this mutation directly on the human isoform of tropomyosin that is involved in nemaline myopathy. It also represents the first time that human tpm3 has been produced using BEVS. This system can now be used to accurately demonstrate the effect of this (and other disease-associated tropomyosin mutations) on the interactions of tpm3 with the other protein components of the muscle thin filament, including those responsible for differing forms of nemaline myopathy.

Original languageEnglish (US)
Pages (from-to)300-304
Number of pages5
JournalBiochemical and Biophysical Research Communications
Volume296
Issue number2
DOIs
StatePublished - 2002

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Biophysics
  • Biochemistry
  • Cell Biology

Keywords

  • Baculovirus
  • Human slow skeletal muscle α tropomyosin
  • Nemaline myopathy
  • TPM3

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