NDLI: Screening for novel lipolytic enzymes from uncultured soil microorganisms

Content Provider	Springer Nature Link
Author	Lee, Seon Woo Won, Keehoon Lim, He Kyoung Kim, Jin Cheol Choi, Gyung Ja Cho, Kwang Yun
Copyright Year	2004
Abstract	The construction and screening of metagenomic libraries constitute a valuable resource for obtaining novel biocatalysts. In this work, we present the construction of a metagenomic library in Escherichia coli using fosmid and microbial DNA directly isolated from forest topsoil and screened for lipolytic enzymes. The library consisted of 33,700 clones with an average DNA insert size of 35 kb. Eight unique lipolytic active clones were obtained from the metagenomic library on the basis of tributyrin hydrolysis. Subsequently, secondary libraries in a high-copy-number plasmid were generated to select lipolytic subclones and to characterize the individual genes responsible for the lipolytic activity. DNA sequence analysis of six genes revealed that the enzymes encoded by the metagenomic genes for lipolytic activity were novel with 34–48% similarity to known enzymes. They had conserved sequences similar to those in the hormone-sensitive lipase family. Based on their deduced amino acid similarity, the six genes encoding lipolytic enzymes were further divided into three subgroups, the identities among which ranged from 33% to 45%. The six predicted gene products were successfully expressed in E. coli and secreted into the culture broth. Most of the secreted enzymes showed a catalytic activity for hydrolysis of p-nitrophenyl butyrate (C4) but not p-nitrophenyl palmitate (C16).
Starting Page	720
Ending Page	726
Page Count	7
File Format	PDF
ISSN	01757598
Journal	Applied Microbiology and Biotechnology
Volume Number	65
Issue Number	6
e-ISSN	14320614
Language	English
Publisher	Springer-Verlag
Publisher Date	2004-09-10
Publisher Place	Berlin, Heidelberg
Access Restriction	Subscribed
Content Type	Text
Resource Type	Article
Subject	Medicine Applied Microbiology and Biotechnology Biotechnology

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