PDBsum entry 5ift

Hydrolase

PDB id: 5ift

Contents

Protein chain

968 a.a.

Ligands

NAG-NAG-BMA-MAN-MAN

NAG-NAG-BMA-MAN-MAN-MAN-MAN-MAN

NAG-NAG-BMA-MAN-MAN-MAN

GLC-GAL

NAG ×6

SO4

DMS ×3

Metals

_CL

Waters ×604

PDB id:

5ift

Name:

Hydrolase

Title:

Structure of e298q-beta-galactosidase from aspergillus niger in complex with 3-b-galactopyranosyl glucose

Structure:

Probable beta-galactosidase a. Chain: a. Synonym: lactase a. Engineered: yes. Mutation: yes

Source:

Aspergillus niger cbs 513.88. Organism_taxid: 425011. Gene: laca, an01g12150. Expressed in: saccharomyces cerevisiae. Expression_system_taxid: 4932.

Resolution:

2.45Å R-factor: 0.152 R-free: 0.193

Authors:

A.Rico-Diaz,M.Ramirez-Escudero,A.Vizoso Vazquez,M.E.Cerdan,M.Becerra, J.Sanz-Aparicio

Key ref:

A.Rico-Díaz et al. (2017). Structural features of Aspergillus niger β-galactosidase define its activity against glycoside linkages. FEBS J, 284, 1815-1829. PubMed id: 28391618 DOI: 10.1111/febs.14083

Date:

26-Feb-16 Release date: 19-Apr-17

Protein chain

A2QAN3  (BGALA_ASPNC) -  Beta-galactosidase A from Aspergillus niger (strain ATCC MYA-4892 / CBS 513.88 / FGSC A1513)

Seq: 1007 a.a.

Struc: 968 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.3.2.1.23 - beta-galactosidase.
• Reaction: Hydrolysis of terminal, non-reducing beta-D-galactose residues in beta-D-galactosides.

DOI no: 10.1111/febs.14083

FEBS J 284:1815-1829 (2017)

PubMed id: 28391618

Structural features of Aspergillus niger β-galactosidase define its activity against glycoside linkages.

A.Rico-Díaz, M.Ramírez-Escudero, �.�.Vizoso-Vázquez, M.E.Cerdán, M.Becerra, J.Sanz-Aparicio.

ABSTRACT

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.