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Expression, Isolation, and Purification of Soluble and Insoluble Biotinylated Proteins for Nerve Tissue Regeneration
Published on: January 22, 2014
Native and heterologous protein secretion by Streptomyces lividans
M Sathyamoorthy1, D Stemke, M K Speedie
1Department of Pharmaceutical Sciences, School of Pharmacy, University of Maryland, Baltimore 21201, USA.
Abstract:
The secretion of the heterologous parathion phosphotriesterase in S. lividans using the Streptomyces beta-galactosidase signal sequence was further characterised using a pulse/chase system. Unsecreted cell-associated protein in both the precursor and signal-cleaved forms was observed when the protein was expressed from both low- and high-copy vectors. Fractionation of the cells followed by immunoprecipitation with phosphotriesterase antibody suggests that the precursor is membrane-bound while the signal cleaved form is present in the soluble fraction. Preliminary data on the processing of alpha-amylase, a native streptomyces protein, showed much more rapid processing and secretion, but nevertheless still revealed cell-associated, signal-cleaved protein.
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