High yield expression of recombinant pro-resilin: lactose-induced fermentation in E. coli and facile purification.
Resilin is an elastic protein with outstanding material properties: high resilience and a very high fatigue lifetime. We are interested in the production of resilin-like proteins which can be photo-chemically cross-linked to form rubbery biomaterials to be used in a variety of industrial and medicinal applications. A method has been developed for producing soluble recombinant proteins in small scale fermentation equipment using glycerol batch for initial growth and primary induction by IPTG at carbon source depletion, followed by new growth in lactose-induced culture. Recombinant rec1-resilin has been over-expressed in the host strain Escherichia coli BL21(DE3)pLysS at a level of up to 300 mg/l, a greater than 20-fold increase in volumetric productivity, relative to that obtained from conventional IPTG induction in LB medium. The primary induction step before lactose induction in fresh medium resulted in a 2.5- to 3-fold increase of both volumetric productivity and cell specific yield compared to that without primary induction under the same conditions. This method is amenable and suitable for large scale production of soluble resilin-like proteins at a low operating cost. In addition, a simple 'salt precipitation and heat purification' method allowed rapid and efficient downstream processing of a large quantity of soluble recombinant resilin-like proteins. These methods will enable investigation of the structural and functional properties of resilin-like proteins, and the development of highly resilient biomaterials.
Kim
Misook
M
CSIRO Livestock Industries, Queensland Bioscience Precinct, St. Lucia 4067, Australia. misook.kim@csiro.au
Elvin
Christopher
C
Brownlee
Alan
A
Lyons
Russell
R
D000818:Animals; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development; D005285:Fermentation; D019476:Insect Proteins / Q000235:genetics* / Q000378:metabolism; D007313:Insecta; D007448:Invertebrates; D007785:Lactose / Q000494:pharmacology*; D011498:Protein Precursors / Q000235:genetics; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism
D019476:Insect Proteins; D011498:Protein Precursors; D011994:Recombinant Proteins; C030951:resilin; D007785:Lactose