Over-production of various secretory-form proteins in Streptomyces lividans.
Streptomyces lividans is known to produce large amounts of proteins in culture supernatants. In this report, to expand the secretory expression system with a strong promoter derived from phospholipase D of Streptoverticillium cinnamoneum, we expressed three kinds of proteins: transglutaminase from Stv. cinnamoneum (StvcMTG) and beta-1,4-endoglucanase and beta-glucosidase from Thermobifida fusca YX. The StvcMTG gene was introduced into S. lividans using the shuttle vector pUC702 for Escherichia coli and S. lividans, and high level secretory production of StvcMTG (230 microg/ml in the culture supernatant) was achieved. The other prokaryotic proteins, beta-1,4-endoglucanase and beta-glucosidase, were also expressed in (His)(6)-tag fused form into culture supernatants and retained high activity. Furthermore, complete purification was achieved by conventional column or affinity column chromatography for each recombinant protein with 1 mg/ml over protein concentration. Three independent proteins were thus successfully expressed and purified, and we expect to use this system for the expression of other valuable heterologous proteins.
Noda
Shuhei
S
Department of Chemical Science and Engineering, Graduate School of Engineering, Kobe University, 1-1 Rokkodaicho, Nada, Kobe 657-8501, Japan.
Ito
Yuko
Y
Shimizu
Nobuaki
N
Tanaka
Tsutomu
T
Ogino
Chiaki
C
Kondo
Akihiko
A
D000192:Actinomycetales / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002480:Cellulase / Q000235:genetics / Q000378:metabolism*; D002846:Chromatography, Affinity; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005822:Genetic Vectors; D006026:Glycoside Hydrolases / Q000235:genetics / Q000378:metabolism*; D006639:Histidine / Q000737:chemistry; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D048170:Streptomyces lividans / Q000235:genetics / Q000378:metabolism*; D011503:Transglutaminases / Q000235:genetics / Q000378:metabolism*; D001617:beta-Glucosidase / Q000235:genetics / Q000378:metabolism*
D001426:Bacterial Proteins; D011994:Recombinant Proteins; D006639:Histidine; D011503:Transglutaminases; D006026:Glycoside Hydrolases; D001617:beta-Glucosidase; D002480:Cellulase; C094192:arabinogalactan endo-1,4-beta-galactosidase