Purification, characterization, and molecular cloning of acidophilic xylanase from penicillium sp.40.
Penicillum sp. 40, which can grow in an extremely acidic medium at pH 2.0 was screened from an acidic soil. This fungus produces xylanases when grown in a medium containing xylan as a sole carbon source. A major xylanase was purified from the culture supernatant of Penicillium sp. 40 and designated XynA. The molecular mass of XynA was estimated to be 25,000 by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. XynA has an optimum pH at 2.0 and is stable in pH 2.0-5.0. Western blot analysis using anit-XynA antibody showed that XynA was induced by xylan and repressed by glucose. Also, its production was increased by an acidic medium. The gene encoding XynA (xynA) was isolated from the genomic library of Penicillium sp. 40. The structural part of xynA was found to be 721 bp. The nucleotide sequence of cDNA amplified by RT-PCR showed that the open reading frame of xynA was interrupted by a single intron which was 58 bp in size and encoded 221 amino acids. Direct N-terminal amino acid sequencing showed that the precursor of XynA had a signal peptide composed of 31 amino acids. The molecular mass caliculated from the deduced amino acid sequence of XynA is 20,713. This is lower than that estimated by gel electrophoresis, suggesting that XynA is a glycoprotein. The predicted amino acid sequence of XynA has strong similarity to other family xylanases from fungi.
Kimura
T
T
Faculty of Bioresources, Mie University, Tsu, Japan. t-kimura@bio.mie-u.ac.jp
Ito
J
J
Kawano
A
A
Makino
T
T
Kondo
H
H
Karita
S
S
Sakka
K
K
Ohmiya
K
K
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics; D043364:Endo-1,4-beta Xylanases; D004795:Enzyme Stability; D005800:Genes, Fungal; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010407:Penicillium / Q000201:enzymology* / Q000235:genetics*; D017386:Sequence Homology, Amino Acid; D014995:Xylosidases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification*
D017931:DNA Primers; D018076:DNA, Complementary; D004271:DNA, Fungal; D014995:Xylosidases; D043364:Endo-1,4-beta Xylanases