Naslov
Production and Overexpression of Streptomyces rimosus GDS(L) Lipase in a Heterologous Host

Autori
Bielen, Ana ; Pigac, Jasenka ; Vujaklija, Dušica

Vrsta, podvrsta i kategorija rada
Sažeci sa skupova, sažetak, znanstveni

Izvornik
Book of Abstracts of the Second Congress of Croatian Geneticists / Franekić Čolić, Jasna ; Ugarković, Đurđica - Zagreb : Hrvatsko genetičko društvo, 2005, 63-63

Skup
Congress of Croatian Geneticists (2 ; 2005)

Mjesto i datum
Supetar, Hrvatska, 22.09.2005. - 27.09.2005

Vrsta sudjelovanja
Poster

Vrsta recenzije
Domaća recenzija

Ključne riječi
lipases; Streptomyces rimosus; Streptomyces lividans

Sažetak
Lipases are currently attracting an enormous biotechnological attention because of their potential to catalyze both hydrolysis and synthesis reactions with high regio- and enantioselectivity. Lipase from Streptomyces rimosus has been previously purified and biochemicaly characterized as an enzyme with significant industrial potential. Streptomyces rimosus lipase gene has been PCR amplified with the 5’ -primer containing original RBS and the 3’ -primer encoding a 6xHis. Lipase was overexpressed using pANT 849 vector in a heterologous host, S. lividans TK23. With this system we avoided previously observed plasmid structural instability. The time course of the lipase production in S. lividans was analysed carefully by quantitative measurements of biomass and lipase activity, detected spectrophotometrically and on agar plates. Lipase activity varied in the culture supernatants from different media and it was growth dependent. Maximum lipase activity was determined in the late stationary phase of growth in GR2d medium. Lipase was purified from supernatant in a three-step procedure. Affinity chromatography with Ni-NTA resin has been used to purify mature lipase with His tagged C-terminus. Introduction of two additional steps: desalting of the proteins in the supernatant by gel-filtration on Sephadex G-25 and binding to CM-cellulose, significantly increased binding of the lipase to the Ni-NTA affinity matrix.

Izvorni jezik
Engleski

Znanstvena područja
Biologija

POVEZANOST RADA