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PDBsum entry 5ift
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References listed in PDB file
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Key reference
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Title
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Structural features of aspergillus niger β-Galactosidase define its activity against glycoside linkages.
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Authors
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A.Rico-Díaz,
M.Ramírez-Escudero,
..Vizoso-Vázquez,
M.E.Cerdán,
M.Becerra,
J.Sanz-Aparicio.
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Ref.
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FEBS J, 2017,
284,
1815-1829.
[DOI no: ]
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PubMed id
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Abstract
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β-Galactosidases are biotechnologically interesting enzymes that catalyze the
hydrolysis or transgalactosylation of β-galactosides. Among them, the
Aspergillus niger β-galactosidase (AnβGal) belongs to the glycoside hydrolase
family 35 (GH35) and is widely used in the industry due to its high hydrolytic
activity which is used to degrade lactose. We present here its three-dimensional
structure in complex with different oligosaccharides, to illustrate the
structural determinants of the broad specificity of the enzyme against different
glycoside linkages. Remarkably, the residues Phe264, Tyr304, and Trp806 make a
dynamic hydrophobic platform that accommodates the sugar at subsite +1
suggesting a main role on the recognition of structurally different substrates.
Moreover, complexes with the trisaccharides show two potential subsites +2
depending on the substrate type. This feature and the peculiar shape of its wide
cavity suggest that AnβGal might accommodate branched substrates from the
complex net of polysaccharides composing the plant material in its natural
environment. Relevant residues were selected and mutagenesis analyses were
performed to evaluate their role in the catalytic performance and the
hydrolase/transferase ratio of AnβGal. Thus, we generated mutants with improved
transgalactosylation activity. In particular, the variant
Y304F/Y355H/N357G/W806F displays a higher level of galacto-oligosaccharides
production than the Aspergillus oryzae β-galactosidase, which is the preferred
enzyme in the industry owing to its high transferase activity. Our results
provide new knowledge on the determinants modulating specificity and the
catalytic performance of fungal GH35 β-galactosidases. In turn, this
fundamental background gives novel tools for the future improvement of these
enzymes, which represent an interesting target for rational design.
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