Unusual microbial xylanases from insect guts.
Recombinant DNA technologies enable the direct isolation and expression of novel genes from biotopes containing complex consortia of uncultured microorganisms. In this study, genomic libraries were constructed from microbial DNA isolated from insect intestinal tracts from the orders Isoptera (termites) and Lepidoptera (moths). Using a targeted functional assay, these environmental DNA libraries were screened for genes that encode proteins with xylanase activity. Several novel xylanase enzymes with unusual primary sequences and novel domains of unknown function were discovered. Phylogenetic analysis demonstrated remarkable distance between the sequences of these enzymes and other known xylanases. Biochemical analysis confirmed that these enzymes are true xylanases, which catalyze the hydrolysis of a variety of substituted beta-1,4-linked xylose oligomeric and polymeric substrates and produce unique hydrolysis products. From detailed polyacrylamide carbohydrate electrophoresis analysis of substrate cleavage patterns, the xylan polymer binding sites of these enzymes are proposed.
Brennan
YaLi
Y
Diversa Corporation, San Diego, CA 92121, USA.
Callen
Walter N
WN
Christoffersen
Leif
L
Dupree
Paul
P
Goubet
Florence
F
Healey
Shaun
S
Hernández
Myrian
M
Keller
Martin
M
Li
Ke
K
Palackal
Nisha
N
Sittenfeld
Ana
A
Tamayo
Giselle
G
Wells
Steve
S
Hazlewood
Geoffrey P
GP
Mathur
Eric J
EJ
Short
Jay M
JM
Robertson
Dan E
DE
Steer
Brian A
BA
D000595:Amino Acid Sequence; D000818:Animals; D001419:Bacteria / Q000201:enzymology* / Q000235:genetics; D004269:DNA, Bacterial / Q000032:analysis / Q000302:isolation & purification; D004271:DNA, Fungal / Q000032:analysis / Q000302:isolation & purification; D004064:Digestive System / Q000382:microbiology*; D043364:Endo-1,4-beta Xylanases / Q000737:chemistry / Q000145:classification / Q000235:genetics* / Q000378:metabolism*; D005658:Fungi / Q000201:enzymology* / Q000235:genetics; D015723:Gene Library; D020049:Isoptera / Q000382:microbiology*; D008969:Molecular Sequence Data; D009036:Moths / Q000382:microbiology*; D010802:Phylogeny; D016415:Sequence Alignment
D004269:DNA, Bacterial; D004271:DNA, Fungal; D043364:Endo-1,4-beta Xylanases