Structural analysis of human skeletal beta-tropomyosin produced in E. coli.
We have cloned the cDNA coding the beta-tropomyosin of human muscle in an expression vector whose expression depends upon a promotor that can be induced by isopropyl-beta-thiogalactopyranoside. We show that a new protein was synthesized by bacteria containing the engineered plasmid. This protein was heat stable and reacted with antibodies against tropomyosin. We have purified this protein and further identified it by determining its amino acid composition and sequencing the NH2 terminal. Unlike the native muscle tropomyosin, the NH2 terminal is not acetylated and contains a methionine. The circular dichroism spectrum is compatible with 100% alpha-helices. These results show that the protein synthesized in E. coli possesses a native structure.
Ferraz
C
C
CRBM du CNRS, U249 INSERM, Montpellier, France.
Derancourt
J
J
Sri Widada
J
J
Liautard
J P
JP
D000595:Amino Acid Sequence; D015153:Blotting, Western; D002942:Circular Dichroism; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D014335:Tropomyosin / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification
D011994:Recombinant Proteins; D014335:Tropomyosin