TY - JOUR
T1 - Modulation of the transglycosylation activity of plant family GH18 chitinase by removing or introducing a tryptophan side chain
AU - Umemoto, Naoyuki
AU - Ohnuma, Takayuki
AU - Osawa, Takuo
AU - Numata, Tomoyuki
AU - Fukamizo, Tamo
N1 - Funding Information:
We thank the beam-line staff at BL-17A of the Photon Factory (Ibaraki, Japan) for technical assistance during the data collection. This work was supported by the Strategic Project to Support the Formation of Research Bases at Private Universities: Matching Fund Subsidy from ‘MEXT’ – Japan (Ministry of Education, Culture, Sports, Science and Technology), 2011–015 ( S1101035 ). NU was supported by a Research Fellowship for Young Scientists from the Japan Society for the Promotion of Science ( 26-7951 ).
Publisher Copyright:
© 2015 Federation of European Biochemical Societies.
PY - 2015/8/12
Y1 - 2015/8/12
N2 - Abstract Transglycosylation (TG) activity of a family GH18 chitinase from the cycad, Cycas revoluta, (CrChiA) was modulated by removing or introducing a tryptophan side chain. The removal from subsite +3 through mutation of Trp168 to alanine suppressed TG activity, while introduction into subsite +1 through mutation of Gly77 to tryptophan (CrChiA-G77W) enhanced TG activity. The crystal structures of an inactive double mutant of CrChiA (CrChiA-G77W/E119Q) with one or two N-acetylglucosamine residues occupying subsites +1 or +1/+2, respectively, revealed that the Trp77 side chain was oriented toward +1 GlcNAc to be stacked with it face-to-face, but rotated away from subsite +1 in the absence of GlcNAc at the subsite. Aromatic residues in the aglycon-binding site are key determinants of TG activity of GH18 chitinases.
AB - Abstract Transglycosylation (TG) activity of a family GH18 chitinase from the cycad, Cycas revoluta, (CrChiA) was modulated by removing or introducing a tryptophan side chain. The removal from subsite +3 through mutation of Trp168 to alanine suppressed TG activity, while introduction into subsite +1 through mutation of Gly77 to tryptophan (CrChiA-G77W) enhanced TG activity. The crystal structures of an inactive double mutant of CrChiA (CrChiA-G77W/E119Q) with one or two N-acetylglucosamine residues occupying subsites +1 or +1/+2, respectively, revealed that the Trp77 side chain was oriented toward +1 GlcNAc to be stacked with it face-to-face, but rotated away from subsite +1 in the absence of GlcNAc at the subsite. Aromatic residues in the aglycon-binding site are key determinants of TG activity of GH18 chitinases.
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U2 - 10.1016/j.febslet.2015.07.018
DO - 10.1016/j.febslet.2015.07.018
M3 - Article
C2 - 26216755
AN - SCOPUS:84938953442
SN - 0014-5793
VL - 589
SP - 2327
EP - 2333
JO - FEBS Letters
JF - FEBS Letters
IS - 18
M1 - 37276
ER -