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The dimeric structure of wild-type human glycosyltransferase B4GalT1

DOI: 10.1371/journal.pone.0205571 DOI Help

Authors: Deborah Harrus (University of Oulu) , Fawzi Khoder-Agha (University of Oulu) , Miika Peltoniemi (University of Oulu) , Antti Hassinen (University of Oulu) , Lloyd Ruddock (University of Oulu) , Sakari Kellokumpu (University of Oulu) , Tuomo Glumoff (University of Oulu)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Plos One , VOL 13

State: Published (Approved)
Published: October 2018
Diamond Proposal Number(s): 14794

Open Access Open Access

Abstract: Most glycosyltransferases, including B4GalT1 (EC, are known to assemble into enzyme homomers and functionally relevant heteromers in vivo. However, it remains unclear why and how these enzymes interact at the molecular/atomic level. Here, we solved the crystal structure of the wild-type human B4GalT1 homodimer. We also show that B4GalT1 exists in a dynamic equilibrium between monomer and dimer, since a purified monomer reappears as a mixture of both and as we obtained crystal forms of the monomer and dimer assemblies in the same crystallization conditions. These two crystal forms revealed the unliganded B4GalT1 in both the open and the closed conformation of the Trp loop and the lid regions, responsible for donor and acceptor substrate binding, respectively. The present structures also show the lid region in full in an open conformation, as well as a new conformation for the GlcNAc acceptor loop (residues 272–288). The physiological relevance of the homodimer in the crystal was validated by targeted mutagenesis studies coupled with FRET assays. These showed that changing key catalytic amino acids impaired homomer formation in vivo. The wild-type human B4GalT1 structure also explains why the variant proteins used for crystallization in earlier studies failed to reveal the homodimers described in this study.

Journal Keywords: Dimers; Crystal structure; Enzyme structure; Monomers; Fluorescence resonance energy transfer; Crystallization; Electron density; Enzymes

Diamond Keywords: Enzymes

Subject Areas: Biology and Bio-materials

Instruments: I04-Macromolecular Crystallography

Added On: 07/11/2018 11:28


Discipline Tags:

Structural biology Life Sciences & Biotech

Technical Tags:

Diffraction Macromolecular Crystallography (MX)