Prospecting for novel biocatalysts in a soil metagenome

被引:169
作者
Voget, S
Leggewie, C
Uesbeck, A
Raasch, C
Jaeger, KE
Streit, WR
机构
[1] Univ Gottingen, Inst Mikrobiol & Genet, D-37077 Gottingen, Germany
[2] Univ Gottingen, Lab Genomanalyse, D-37077 Gottingen, Germany
[3] Univ Dusseldorf, Inst Mol Enzymtechnol, Forschungszentrum Julich, D-52425 Julich, Germany
关键词
D O I
10.1128/AEM.69.10.6235-6242.2003
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
The metagenomes of complex microbial communities are rich sources of novel biocatalysts. We exploited the metagenome of a mixed microbial population for isolation of more than 15 different genes encoding novel biocatalysts by using a combined cultivation and direct cloning strategy. A 16S rRNA sequence analysis revealed the presence of hitherto uncultured microbes closely related to the genera Pseudomonas, Agrobacterium, Xanthomonas, Microbulbifer, and Janthinobacterium. Total genomic DNA from this bacterial community was used to construct cosmid DNA libraries, which were functionally searched for novel enzymes of biotechnological value. Our searches in combination with cosmid sequencing resulted in identification of four clones encoding 12 putative agarase genes, most of which were organized in clusters consisting of two or three genes. Interestingly, nine of these agarase genes probably originated from gene duplications. Furthermore, we identified by DNA sequencing several other biocatalyst-encoding genes, including genes encoding a putative stereoselective amidase (ami4), two cellulases (gnuB and uvs080), an alpha-amylase (amyA) a 1,4-alpha-glucan branching enzyme (amyB), and two pectate lyases (pelA and uvs119). Also, a conserved cluster of two lipase genes was identified, which was linked to genes encoding a type I secretion system. The novel gene aguB was overexpressed in Escherichia coli, and the enzyme activities were determined. Finally, we describe more than 162 kb of DNA sequence that provides a strong platform for further characterization of this microbial consortium.
引用
收藏
页码:6235 / 6242
页数:8
相关论文
共 33 条
  • [1] Industrial applications of pectic enzymes: a review
    Alkorta, I
    Garbisu, C
    Llama, MJ
    Serra, JL
    [J]. PROCESS BIOCHEMISTRY, 1998, 33 (01) : 21 - 28
  • [2] PHYLOGENETIC IDENTIFICATION AND IN-SITU DETECTION OF INDIVIDUAL MICROBIAL-CELLS WITHOUT CULTIVATION
    AMANN, RI
    LUDWIG, W
    SCHLEIFER, KH
    [J]. MICROBIOLOGICAL REVIEWS, 1995, 59 (01) : 143 - 169
  • [3] Construction and analysis of bacterial artificial chromosome libraries from a marine microbial assemblage
    Béjà, O
    Suzuki, MT
    Koonin, EV
    Aravind, L
    Hadd, A
    Nguyen, LP
    Villacorta, R
    Amjadi, M
    Garrigues, C
    Jovanovich, SB
    Feldman, RA
    DeLong, EF
    [J]. ENVIRONMENTAL MICROBIOLOGY, 2000, 2 (05) : 516 - 529
  • [4] Caldwell D., 1997, MANUAL ENV MICROBIOL, P79
  • [5] Phase-variable expression of an operon encoding extracellular alkaline protease, a serine protease homolog, and lipase in Pseudomonas brassicacearum
    Chabeaud, P
    de Groot, A
    Bitter, W
    Tommassen, J
    Heulin, T
    Achouak, W
    [J]. JOURNAL OF BACTERIOLOGY, 2001, 183 (06) : 2117 - 2120
  • [6] THE PSEUDOMONAS-FLUORESCENS LIPASE HAS A C-TERMINAL SECRETION SIGNAL AND IS SECRETED BY A 3-COMPONENT BACTERIAL ABC-EXPORTER SYSTEM
    DUONG, F
    SOSCIA, C
    LAZDUNSKI, A
    MURGIER, M
    [J]. MOLECULAR MICROBIOLOGY, 1994, 11 (06) : 1117 - 1126
  • [7] Direct cloning from enrichment cultures, a reliable strategy for isolation of complete operons and genes from microbial consortia
    Entcheva, P
    Liebl, W
    Johann, A
    Hartsch, T
    Streit, WR
    [J]. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 2001, 67 (01) : 89 - 99
  • [8] HEALY FG, 1995, APPL MICROBIOL BIOT, V43, P667, DOI [10.1007/BF00164771, 10.1007/s002530050468]
  • [9] Henne A, 1999, APPL ENVIRON MICROB, V65, P3901
  • [10] Screening of environmental DNA libraries for the presence of genes conferring lipolytic activity on Escherichia coli
    Henne, A
    Schmitz, RA
    Bömeke, M
    Gottschalk, G
    Daniel, R
    [J]. APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 2000, 66 (07) : 3113 - 3116