Isolation of an alkane-degrading Alcanivorax sp. strain 2B5 and cloning of the alkB gene.
A Gram-negative coccus, strain 2B5, was isolated from the sea mud of the crude oil-polluted Donghai area in China and identified as Alcanivorax sp. based on its physiological characteristics and analysis of its 16S rRNA gene sequence. Strain 2B5 was able to degrade C(13)-C(30)n-alkanes and branched alkanes (pristane and phytane) from crude oil as the sole carbon source. The optimal temperature and pH for strain 2B5 growth and octadecane degradation were 30-37 degrees C and pH 6.0-7.0, respectively. NaCl is required for strain 2B5 growth and octadecane degradation were highest at NaCl concentrations of 20 to 90 g L(-1). A novel alkane hydroxylase gene (alkB), obtained by self-formed adaptor PCR (SEFA-PCR), displays 41.9% deduced amino acid sequence identity with alkB1 of Alcanivorax borkumensis SK2. Functional heterologous expression of the alkB gene was achieved in Pseudomonas fluorescens KOB2delta1. Recombinant Pseudomonas containing the alkB gene of Alcanivorax sp. strain 2B5 recovered the ability to grow on C(14) and C(16)n-alkanes. RT-PCR analysis showed that expression of the alkB gene was induced by octadecane. There could be other alkane hydroxylases in Alcanivorax sp. 2B5 that could be involved in the degradation of short- and long-chain alkanes.
Liu
Yi-Chen
YC
Key Laboratory of Microbiological Engineering of Agricultural Environment, Ministry of Agriculture, Nanjing Agricultural University, Nanjing 210095, China.
Li
Ling-Zhi
LZ
Wu
Ying
Y
Tian
Wei
W
Zhang
Li-Ping
LP
Xu
Lian
L
Shen
Qi-Rong
QR
Shen
Biao
B
D053521:Alcanivoraceae / Q000145:classification / Q000302:isolation & purification* / Q000502:physiology*; D000473:Alkanes / Q000378:metabolism*; D003001:Cloning, Molecular; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism*; D006899:Mixed Function Oxygenases / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism*; D013045:Species Specificity
D000473:Alkanes; D029968:Escherichia coli Proteins; D011994:Recombinant Proteins; D006899:Mixed Function Oxygenases; C466427:AlkB protein, E coli