High Resolution Structures of the Human ABO(H) Blood Group Enzymes in Complex with Donor Analogs Reveal That the Enzymes Utilize Multiple Donor Conformations to Bind Substrates in a Stepwise Manner

Gagnon, S.; Meloncelli, Peter J.; Zheng, Ruixiang Blake; Haji-Ghassemi, O.; Johal, A.R.; Borisova, S.N.; Lowary, Todd L.; Evans, Stephen V.

doi:10.1074/jbc.m115.682401

Cited by 14 publications

(11 citation statements)

References 48 publications

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“…It is interesting to note that ADP in chain B of the CLg1GBSS structure adopts a conformation quite similar to that observed in structures of some GTA-fold glycosyltransferases which have donors with a covalently bound glucosyl group, for example R. marinus MGS (PDB_ID 2bo8 and 2y4m) ( Flint et al, 2005 ; Nielsen et al, 2011 ) and the human blood group glycosyltransferase (PDB_IDs 5c3b and 5c1l) ( Gagnon et al, 2015 ).…”

Section: Discussionsupporting

confidence: 53%

Crystal Structures of the Catalytic Domain of Arabidopsis thaliana Starch Synthase IV, of Granule Bound Starch Synthase From CLg1 and of Granule Bound Starch Synthase I of Cyanophora paradoxa Illustrate Substrate Recognition in Starch Synthases

et al. 2018

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Starch synthases (SSs) are responsible for depositing the majority of glucoses in starch. Structural knowledge on these enzymes that is available from the crystal structures of rice granule bound starch synthase (GBSS) and barley SSI provides incomplete information on substrate binding and active site architecture. Here we report the crystal structures of the catalytic domains of SSIV from Arabidopsis thaliana, of GBSS from the cyanobacterium CLg1 and GBSSI from the glaucophyte Cyanophora paradoxa, with all three bound to ADP and the inhibitor acarbose. The SSIV structure illustrates in detail the modes of binding for both donor and acceptor in a plant SS. CLg1GBSS contains, in the same crystal structure, examples of molecules with and without bound acceptor, which illustrates the conformational changes induced upon acceptor binding that presumably precede catalytic activity. With structures available from several isoforms of plant and non-plant SSs, as well as the closely related bacterial glycogen synthases, we analyze, at the structural level, the common elements that define a SS, the elements that are necessary for substrate binding and singularities of the GBSS family that could underlie its processivity. While the phylogeny of the SSIII/IV/V has been recently discussed, we now further report the detailed evolutionary history of the GBSS/SSI/SSII type of SSs enlightening the origin of the GBSS enzymes used in our structural analysis.

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Section: Discussionsupporting

confidence: 53%

Crystal Structures of the Catalytic Domain of Arabidopsis thaliana Starch Synthase IV, of Granule Bound Starch Synthase From CLg1 and of Granule Bound Starch Synthase I of Cyanophora paradoxa Illustrate Substrate Recognition in Starch Synthases

et al. 2018

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“…However, the basis of these allosteric interactions is not understood yet. Interestingly, a recent crystallographic study shows that GTA, GTB, and mutants are able to bind to a variety of conformations of donor substrates, including low‐energy solution conformations . It is suggested that the enzymes shift the donor substrates into the catalytically competent “tucked‐under” conformation in a stepwise fashion.…”

Section: Resultsmentioning

confidence: 99%

Protein NMR Studies of Substrate Binding to Human Blood Group A and B Glycosyltransferases

et al. 2017

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Donor and acceptor substrate binding to human blood group A and B glycosyltransferases (GTA, GTB) has been studied by a variety of protein NMR experiments. Prior crystallographic studies had shown these enzymes to adopt an open conformation in the absence of substrates. Binding either of the donor substrate UDP‐Gal or of UDP induces a semiclosed conformation. In the presence of both donor and acceptor substrates, the enzymes shift towards a closed conformation with ordering of an internal loop and the C‐terminal residues, which then completely cover the donor‐binding pocket. Chemical‐shift titrations of uniformly 2H,15N‐labeled GTA or GTB with UDP affected about 20 % of all crosspeaks in 1H,15N TROSY‐HSQC spectra, reflecting substantial plasticity of the enzymes. On the other hand, it is this conformational flexibility that impedes NH backbone assignments. Chemical‐shift‐perturbation experiments with δ1‐[13C]methyl‐Ile‐labeled samples revealed two Ile residues—Ile123 at the bottom of the UDP binding pocket, and Ile192 as part of the internal loop—that were significantly disturbed upon stepwise addition of UDP and H‐disaccharide, also revealing long‐range perturbations. Finally, methyl TROSY‐based relaxation dispersion experiments do not reveal micro‐ to millisecond timescale motions. Although this study reveals substantial conformational plasticity of GTA and GTB, the matter of how binding of substrates shifts the enzymes into catalytically competent states remains enigmatic.

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“…It has four different phenotypes-A, B, AB, and O-that differ in the carbohydrate antigenst hat are dominantly presented on the cell wall of erythrocytes. [2][3][4][5] Even thought he ABO blood group system has been known for more than 100 years and can be routinelys erotyped, its biological functionr emains elusive. [1] In the Aa nd Bp henotype, the Ha ntigen is modified by the addition of N-acetylgalactosamine (A)o rg alactose (B) that are each transferred by the action of GTAo rG TB, respectively.…”

Section: Introductionmentioning

confidence: 99%

“…The mechanistic basis of their specificity has therefore been studied intensively. [2][3][4][5] Even thought he ABO blood group system has been known for more than 100 years and can be routinelys erotyped, its biological functionr emains elusive. [1] However,alarge body of work has linked blood subgroups, in particularn on-O phenotypes, to the incidence of various cancer types and their progression, although no underlying biological mechanism has been establishedf or this yet.…”

Section: Introductionmentioning

confidence: 99%

Fragment Growing to Design Optimized Inhibitors for Human Blood Group B Galactosyltransferase (GTB)

et al. 2019

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Human blood group B galactosyltransferase (GTB) catalyzes the galactosylation of the H antigen and is responsible for the formation of the blood group antigen of phenotype B. The ABO blood group system is well studied and routinely serotyped before transfusion and transplantation. Blood type subgroups have been repeatedly linked to an increased occurrence of diseases (e.g., a highly increased incidence rate for pancreatic cancer for individuals with blood group phenotype B). 3‐Phenyl‐5‐(piperazin‐1‐yl)‐1,2,4‐thiadiazole 1 has previously been described to inhibit GTB with a Ki value of 800 μm. In this work, we describe a computer‐guided fragment‐growing approach for the optimization of this fragment that was subsequently realized by synthesizing the most promising ligands. Enlarging the phenyl moiety of fragment 1 to a naphthyl moiety resulted in ligand 3‐(naphthalene‐1‐yl)‐5‐(piperazin‐1‐yl)‐1,2,4‐thiadiazole 2 a, which shows a threefold improvement in binding affinity (Ki=271 μm).

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High Resolution Structures of the Human ABO(H) Blood Group Enzymes in Complex with Donor Analogs Reveal That the Enzymes Utilize Multiple Donor Conformations to Bind Substrates in a Stepwise Manner

Cited by 14 publications

References 48 publications

Crystal Structures of the Catalytic Domain of Arabidopsis thaliana Starch Synthase IV, of Granule Bound Starch Synthase From CLg1 and of Granule Bound Starch Synthase I of Cyanophora paradoxa Illustrate Substrate Recognition in Starch Synthases

Crystal Structures of the Catalytic Domain of Arabidopsis thaliana Starch Synthase IV, of Granule Bound Starch Synthase From CLg1 and of Granule Bound Starch Synthase I of Cyanophora paradoxa Illustrate Substrate Recognition in Starch Synthases

Protein NMR Studies of Substrate Binding to Human Blood Group A and B Glycosyltransferases

Fragment Growing to Design Optimized Inhibitors for Human Blood Group B Galactosyltransferase (GTB)

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