Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Crystal structures of human pancreatic alpha-amylase in complex with carbohydrate and proteinaceous inhibitors.

ABSTRACT: Crystal structures of human pancreatic alpha-amylase (HPA) in complex with naturally occurring inhibitors have been solved. The tetrasaccharide acarbose and a pseudo-pentasaccharide of the trestatin family produced identical continuous electron densities corresponding to a pentasaccharide species, spanning the -3 to +2 subsites of the enzyme, presumably resulting from transglycosylation. Binding of the acarviosine core linked to a glucose residue at subsites -1 to +2 appears to be a critical part of the interaction process between alpha-amylases and trestatin-derived inhibitors. Two crystal forms, obtained at different values of pH, for the complex of HPA with the protein inhibitor from Phaseolus vulgaris (alpha-amylase inhibitor) have been solved. The flexible loop typical of the mammalian alpha-amylases was shown to exist in two different conformations, suggesting that loop closure is pH-sensitive. Structural information is provided for the important inhibitor residue, Arg-74, which has not been observed previously in structural analyses.

SUBMITTER: Nahoum V

PROVIDER: S-EPMC1220841 | biostudies-other | 2000 Feb

REPOSITORIES: biostudies-other

ACCESS DATA

Json Xml

Similar Datasets

Structures of two novel crystal forms of Aspergillus oryzae alpha amylase (taka-amylase).

Project description:The structures of Aspergillus oryzae α-amylase were determined in a tetragonal crystal, having one molecule as asymmetric unit, and a monoclinic crystal with two molecules as asymmetric unit. Both crystal forms were obtained from trace contaminants of an old commercial lipase preparation. Structures were determined and refined to 1.65 Å and 1.43 Å resolution respectively. The latter crystal has a non-crystallographic (NCS) twofold axis within the asymmetric unit. Glycosylation at Asn197 is evident, and in the tetragonal crystal can be seen to include three, partially disordered sugar residues following the initial N-acetyl glucosamine (NAG). Superposition of the tetragonal crystal model on the α-amylases from Bacillus subtilis (PDB:1BAG), pig pancreas (PDB:3L2L), and barley (PDB:1AMY), show a high degree of coincidence, particularly for the (β/α)8-barrel domains, and especially within the active site. Using this structural agreement between amylases, we extrapolated the binding model of a six residue, limit dextrin found in pig pancreas α-amylase to the A. oryzae enzyme model, which predicts substrate interacting amino acid residues.

| S-EPMC8187280 | biostudies-literature

Synthesis of montbretin A analogues yields potent competitive inhibitors of human pancreatic α-amylase.

Project description:Simplified analogues of the potent human amylase inhibitor montbretin A were synthesised and shown to bind tightly, K I = 60 and 70 nM, with improved specificity over medically relevant glycosidases, making them promising candidates for controlling blood glucose. Crystallographic analysis confirmed similar binding modes and identified new active site interactions.

| S-EPMC7069248 | biostudies-literature

Crystal structure of Bacillus subtilis alpha-amylase in complex with acarbose.

Project description:The crystal structure of Bacillus subtilis alpha-amylase, in complex with the pseudotetrasaccharide inhibitor acarbose, revealed an hexasaccharide in the active site as a result of transglycosylation. After comparison with the known structure of the catalytic-site mutant complexed with the native substrate maltopentaose, it is suggested that the present structure represents a mimic intermediate in the initial stage of the catalytic process.

| S-EPMC262715 | biostudies-literature

Folding Then Binding vs Folding Through Binding in Macrocyclic Peptide Inhibitors of Human Pancreatic α-Amylase.

Project description:De novo macrocyclic peptides, derived using selection technologies such as phage and mRNA display, present unique and unexpected solutions to challenging biological problems. This is due in part to their unusual folds, which are able to present side chains in ways not available to canonical structures such as α-helices and β-sheets. Despite much recent interest in these molecules, their folding and binding behavior remains poorly characterized. In this work, we present cocrystallization, docking, and solution NMR structures of three de novo macrocyclic peptides that all bind as competitive inhibitors with single-digit nanomolar Ki to the active site of human pancreatic α-amylase. We show that a short stably folded motif in one of these is nucleated by internal hydrophobic interactions in an otherwise dynamic conformation in solution. Comparison of the solution structures with a target-bound structure from docking indicates that stabilization of the bound conformation is provided through interactions with the target protein after binding. These three structures also reveal a surprising functional convergence to present a motif of a single arginine sandwiched between two aromatic residues in the interactions of the peptide with the key catalytic residues of the enzyme, despite little to no other structural homology. Our results suggest that intramolecular hydrophobic interactions are important for priming binding of small macrocyclic peptides to their target and that high rigidity is not necessary for high affinity.

| S-EPMC6700688 | biostudies-literature

Crystal structures of ?-primeverosidase in complex with disaccharide amidine inhibitors.

Project description:?-Primeverosidase (PD) is a disaccharide-specific ?-glycosidase in tea leaves. This enzyme is involved in aroma formation during the manufacturing process of oolong tea and black tea. PD hydrolyzes ?-primeveroside (6-O-?-d-xylopyranosyl-?-d-glucopyranoside) at the ?-glycosidic bond of primeverose to aglycone, and releases aromatic alcoholic volatiles of aglycones. PD only accepts primeverose as the glycone substrate, but broadly accepts various aglycones, including 2-phenylethanol, benzyl alcohol, linalool, and geraniol. We determined the crystal structure of PD complexes using highly specific disaccharide amidine inhibitors, N-?-primeverosylamidines, and revealed the architecture of the active site responsible for substrate specificity. We identified three subsites in the active site: subsite -2 specific for 6-O-?-d-xylopyranosyl, subsite -1 well conserved among ?-glucosidases and specific for ?-d-glucopyranosyl, and wide subsite +1 for hydrophobic aglycone. Glu-470, Ser-473, and Gln-477 act as the specific hydrogen bond donors for 6-O-?-d-xylopyranosyl in subsite -2. On the other hand, subsite +1 was a large hydrophobic cavity that accommodates various aromatic aglycones. Compared with aglycone-specific ?-glucosidases of the glycoside hydrolase family 1, PD lacks the Trp crucial for aglycone recognition, and the resultant large cavity accepts aglycone and 6-O-?-d-xylopyranosyl together. PD recognizes the ?-primeverosides in subsites -1 and -2 by hydrogen bonds, whereas the large subsite +1 loosely accommodates various aglycones. The glycone-specific activity of PD for broad aglycone substrates results in selective and multiple release of temporally stored alcoholic volatile aglycones of ?-primeveroside.

| S-EPMC4059125 | biostudies-literature

Structural and mechanistic studies of chloride induced activation of human pancreatic alpha-amylase.

Project description:The mechanism of allosteric activation of alpha-amylase by chloride has been studied through structural and kinetic experiments focusing on the chloride-dependent N298S variant of human pancreatic alpha-amylase (HPA) and a chloride-independent TAKA-amylase. Kinetic analysis of the HPA variant clearly demonstrates the pronounced activating effect of chloride ion binding on reaction rates and its effect on the pH-dependence of catalysis. Structural alterations observed in the N298S variant upon chloride ion binding suggest that the chloride ion plays a variety of roles that serve to promote catalysis. One of these is having a strong influence on the positioning of the acid/base catalyst residue E233. Absence of chloride ion results in multiple conformations for this residue and unexpected enzymatic products. Chloride ion and N298 also appear to stabilize a helical region of polypeptide chain from which projects the flexible substrate binding loop unique to chloride-dependent alpha-amylases. This structural feature also serves to properly orient the catalytically essential residue D300. Comparative analyses show that the chloride-independent alpha-amylases compensate for the absence of bound chloride by substituting a hydrophobic core, altering the manner in which substrate interactions are made and shifting the placement of N298. These evolutionary differences presumably arise in response to alternative operating environments or the advantage gained in a particular product profile. Attempts to engineer chloride-dependence into the chloride-independent TAKA-amylase point out the complexity of this system, and the fact that a multitude of factors play a role in binding chloride ion in the chloride-dependent alpha-amylases.

| S-EPMC2279296 | biostudies-literature

Crystal structures of three bicyclic carbohydrate derivatives.

Project description:The title compounds, [(1R,3R,4R,5R,6S)-4,5-bis-(acet-yloxy)-7-oxo-2-oxabi-cyclo[4.2.0]octan-3-yl]methyl acetate, C14H18O8, (I), [(1S,4R,5S,6R)-5-acet-yloxy-7-hy-droxy-imino-2-oxobi-cyclo-[4.2.0]octan-4-yl acetate, C11H15NO6, (II), and [(3aR,5R,6R,7R,7aS)-6,7-bis-(acet-yloxy)-2-oxo-octa-hydro-pyrano[3,2-b]pyrrol-5-yl]methyl acetate, C14H19NO8, (III), are stable bicyclic carbohydrate derivatives. They can easily be synthesized in a few steps from commercially available glycals. As a result of the ring strain from the four-membered rings in (I) and (II), the conformations of the carbohydrates deviate strongly from the ideal chair form. Compound (II) occurs in the boat form. In the five-membered lactam (III), on the other hand, the carbohydrate adopts an almost ideal chair conformation. As a result of the distortion of the sugar rings, the configurations of the three bicyclic carbohydrate derivatives could not be determined from their NMR coupling constants. From our three crystal structure determinations, we were able to establish for the first time the absolute configurations of all new stereocenters of the carbohydrate rings.

| S-EPMC5137623 | biostudies-literature

Crystal structures of cholesteryl ester transfer protein in complex with inhibitors.

Project description:Human plasma cholesteryl ester transfer protein (CETP) transports cholesteryl ester from the antiatherogenic high-density lipoproteins (HDL) to the proatherogenic low-density and very low-density lipoproteins (LDL and VLDL). Inhibition of CETP has been shown to raise human plasma HDL cholesterol (HDL-C) levels and is potentially a novel approach for the prevention of cardiovascular diseases. Here, we report the crystal structures of CETP in complex with torcetrapib, a CETP inhibitor that has been tested in phase 3 clinical trials, and compound 2, an analog from a structurally distinct inhibitor series. In both crystal structures, the inhibitors are buried deeply within the protein, shifting the bound cholesteryl ester in the N-terminal pocket of the long hydrophobic tunnel and displacing the phospholipid from that pocket. The lipids in the C-terminal pocket of the hydrophobic tunnel remain unchanged. The inhibitors are positioned near the narrowing neck of the hydrophobic tunnel of CETP and thus block the connection between the N- and C-terminal pockets. These structures illuminate the unusual inhibition mechanism of these compounds and support the tunnel mechanism for neutral lipid transfer by CETP. These highly lipophilic inhibitors bind mainly through extensive hydrophobic interactions with the protein and the shifted cholesteryl ester molecule. However, polar residues, such as Ser-230 and His-232, are also found in the inhibitor binding site. An enhanced understanding of the inhibitor binding site may provide opportunities to design novel CETP inhibitors possessing more drug-like physical properties, distinct modes of action, or alternative pharmacological profiles.

| S-EPMC3481329 | biostudies-literature

Monoclinic crystal form of Aspergillus niger alpha-amylase in complex with maltose at 1.8 angstroms resolution.

Project description:Aspergillus niger alpha-amylase catalyses the hydrolysis of alpha-1,4-glucosidic bonds in starch. It shows 100% sequence identity to the A. oryzae homologue (also called TAKA-amylase), three crystal structures of which have been published to date. Two of them belong to the orthorhombic space group P2(1)2(1)2(1) with one molecule per asymmetric unit and one belongs to the monoclinic space group P2(1) with three molecules per asymmetric unit. Here, the purification, crystallization and structure determination of A. niger alpha-amylase crystallized in the monoclinic space group P2(1) with two molecules per asymmetric unit in complex with maltose at 1.8 angstroms resolution is reported. Furthermore, a novel 1.6 angstroms resolution orthorhombic crystal form (space group P2(1)2(1)2) of the native enzyme is presented. Four maltose molecules are observed in the maltose-alpha-amylase complex. Three of these occupy active-site subsites -2 and -1, +1 and +2 and the hitherto unobserved subsites +4 (Asp233, Gly234) and +5 (Asp235). The fourth maltose molecule binds at the distant binding sites d1 (Tyr382) and d2 (Trp385), also previously unobserved. Furthermore, it is shown that the active-site groove permits different binding modes of sugar units at subsites +1 and +2. This flexibility of the active-site cleft close to the catalytic centre might be needed for a productive binding of substrate chains and/or release of products.

| S-EPMC2242925 | biostudies-literature

Crystal structures of Leishmania mexicana arginase complexed with α,α-disubstituted boronic amino-acid inhibitors.

Project description:Leishmania arginase is a potential drug target for the treatment of leishmaniasis because this binuclear manganese metalloenzyme initiates de novo polyamine biosynthesis by catalyzing the hydrolysis of L-arginine to generate L-ornithine and urea. The product L-ornithine subsequently undergoes decarboxylation to yield putrescine, which in turn is utilized for spermidine biosynthesis. Polyamines such as spermidine are essential for the growth and survival of the parasite, so inhibition of enzymes in the polyamine-biosynthetic pathway comprises an effective strategy for treating parasitic infections. To this end, two X-ray crystal structures of L. mexicana arginase complexed with α,α-disubstituted boronic amino-acid inhibitors based on the molecular scaffold of 2-(S)-amino-6-boronohexanoic acid are now reported. Structural comparisons with human and parasitic arginase complexes reveal interesting differences in the binding modes of the additional α-substituents, i.e. the D side chains, of these inhibitors. Subtle differences in the three-dimensional contours of the outer active-site rims among arginases from different species lead to different conformations of the D side chains and thus different inhibitor-affinity trends. The structures suggest that it is possible to maintain affinity while fine-tuning intermolecular interactions of the D side chain of α,α-disubstituted boronic amino-acid inhibitors in the search for isozyme-specific and species-specific arginase inhibitors.

| S-EPMC4822987 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data