Crystal structures of eukaryote glycosyltransferases reveal biologically relevant enzyme homooligomers

Harrus, Déborah; Kellokumpu, Sakari; Glumoff, Tuomo

doi:10.1007/s00018-017-2659-x

Cited by 19 publications

(15 citation statements)

References 84 publications

(103 reference statements)

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“…Gel filtration analysis indicates that the GnT-V luminal domain and mini-GnT-V likely exist as a monomer in solution (Supplementary Table 3 ). Glycosyltransferases often form homo-oligomers 60 . GnT-V may oligomerize via the stem region and not the catalytic domain (Supplementary Figure 12A ).…”

Section: Discussionmentioning

confidence: 99%

Structure and mechanism of cancer-associated N-acetylglucosaminyltransferase-V

et al. 2018

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N-acetylglucosaminyltransferase-V (GnT-V) alters the structure of specific N-glycans by modifying α1-6-linked mannose with a β1-6-linked N-acetylglucosamine branch. β1-6 branch formation on cell surface receptors accelerates cancer metastasis, making GnT-V a promising target for drug development. However, the molecular basis of GnT-V’s catalytic mechanism and substrate specificity are not fully understood. Here, we report crystal structures of human GnT-V luminal domain with a substrate analog. GnT-V luminal domain is composed of a GT-B fold and two accessary domains. Interestingly, two aromatic rings sandwich the α1-6 branch of the acceptor N-glycan and restrain the global conformation, partly explaining the fine branch specificity of GnT-V. In addition, interaction of the substrate N-glycoprotein with GnT-V likely contributes to protein-selective and site-specific glycan modification. In summary, the acceptor-GnT-V complex structure suggests a catalytic mechanism, explains the previously observed inhibition of GnT-V by branching enzyme GnT-III, and provides a basis for the rational design of drugs targeting N-glycan branching.

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Section: Discussionmentioning

confidence: 99%

Structure and mechanism of cancer-associated N-acetylglucosaminyltransferase-V

et al. 2018

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“…In addition, B4GALT1 has another interaction site for ST3GAL3 and for lactalbumin that are distinct from the ones described in this report. 3 From the functional point of view, the formation of B4GALT1 homomers via the active site interaction surface serves to keep the enzyme mini-mally active until it reaches the Golgi compartment, where its binds ST6GAL1 and exposes its active site because of disassembly of the homodimers. In contrast, the ST6GAL1 enzyme is activated only after it reaches the more oxidizing environment of the Golgi lumen, a condition that is necessary for the formation of two surface-exposed disulfide bonds in the enzyme's catalytic domain.…”

Section: Molecular Interactions Between B4galt1 and St6gal1mentioning

confidence: 99%

“…Most glycosyltransferases are type II transmembrane pro-teins with three distinct structural and functional domains: a transmembrane domain preceded by an N-terminal cytoplasmic tail, a stem domain (STEM) oriented toward the lumen of the Golgi, and a globular catalytic domain (CAT) (2). The presence of the catalytic domains in the Golgi lumen allows for the processing of glycan chains along their passage through the secretory pathway (3).…”

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confidence: 99%

Assembly of B4GALT1/ST6GAL1 heteromers in the Golgi membranes involves lateral interactions via highly charged surface domains

Khoder-Agha

Harrus

Brysbaert

et al. 2019

Journal of Biological Chemistry

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“…Evolutionary conservation of the interface was assessed using the InterEvol server [ 31 ]. The jsPISA score was computed as described in [ 32 ].…”

Section: Methodsmentioning

confidence: 99%

The dimeric structure of wild-type human glycosyltransferase B4GalT1

et al. 2018

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Most glycosyltransferases, including B4GalT1 (EC 2.4.1.38), 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.

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Crystal structures of eukaryote glycosyltransferases reveal biologically relevant enzyme homooligomers

Cited by 19 publications

References 84 publications

Structure and mechanism of cancer-associated N-acetylglucosaminyltransferase-V

Structure and mechanism of cancer-associated N-acetylglucosaminyltransferase-V

Assembly of B4GALT1/ST6GAL1 heteromers in the Golgi membranes involves lateral interactions via highly charged surface domains

The dimeric structure of wild-type human glycosyltransferase B4GalT1

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