Production of human skeletal α-actin proteins by the baculovirus expression system

P.A. Akkari, Kristen Nowak, S.K. Beckman, Kendall Walker, F. Schachat, Nigel Laing

Research output: Contribution to journalArticlepeer-review

15 Citations (Scopus)

Abstract

Mutations within the human skeletal muscle alpha-actin gene cause three different skeletal muscle diseases. Functional studies of the mutant proteins are necessary to better understand the pathogenesis of these diseases, however, no satisfactory system for the expression of mutant muscle actin proteins has been available. We investigated the baculovirus expression vector system (BEVS) for the abundant production of both normal and mutant skeletal muscle alpha-actin. We show that non-mutated actin produced in the BEVS behaves similarly to native actin, as shown by DNase I affinity purification, Western blotting, and consecutive cycles of polymerisation and depolymerisation. Additionally, we demonstrate the production of mutant actin proteins in the BEVS, without detriment to the insect cells in which they are expressed. The BEVS therefore is the method of choice for studying mutant actin proteins causing human diseases.

Original languageEnglish
Pages (from-to)74-79
Number of pages6
JournalBiochemical and Biophysical Research Communications
Volume307
Issue number1
DOIs
Publication statusPublished - 18 Jul 2003

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