@article{3053650,
    title = "Structures of chitobiase mutants complexed with the substrate
di-N-acetyl-D-glucosamine: The catalytic role of the conserved acidic
pair, aspartate 539 and glutamate 540",
    author = "Prag, G and Papanikolau, Y and Tavlas, G and Vorgias, CE and Petratos, K and and Oppenheim, AB",
    journal = "JOURNAL OF MOLECULAR BIOLOGY",
    year = "2000",
    volume = "300",
    number = "3",
    pages = "611-617",
    publisher = "ACADEMIC PRESS LTD- ELSEVIER SCIENCE LTD",
    issn = "0022-2836",
    doi = "10.1006/jmbi.2000.3906",
    keywords = "Serratia marcescens; crystal structure; hexosaminidase;
beta-N-acetylhexosaminidase; catalytic mechanism",
    abstract = "The catalytic domain of chitobiase (beta-N-1-4 acetylhexosaminidase)
from Serratia marcescens, is an alpha/beta TIM-barrel. This enzyme
belongs to family 20 of glycosyl hydrolases in which a conserved amino
acid pair, aspartate-glutamate, is present (Asp539-Glu540). It was
proposed that catalysis by this enzyme family is carried out by
glutamate 540 acting as a proton donor and by the acetamido group of the
substrate as a nucleophile. We investigated the role of Asp539 and
Glu540 by site-directed mutagenesis, biochemical characterization and by
structural analyses of chitobiase substrate co-crystals. We found that
both residues are essential for chitoblase activity. The mutations,
however, led to subtle changes in the catalytic site. Our results
support the model that Glu540 acts as the proton donor and that Asp539
acts in several different ways. Asp539 restrains the acetamido group of
the substrate in a specific orientation by forming a hydrogen bond with
N2 of the non-reduced (-1) sugar. In addition, this residue participates
in substrate binding. It is also required for the correct positioning of
Glu540 and may provide additional negative charge at the active site.
Thus, these biochemical and structural studies provide a molecular
explanation for the functional importance and conservation of these
residues. (C) 2000 Academic Press."
}