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001-es BibID:BIBFORM085848
Első szerző:Gyémánt Gyöngyi (vegyész)
Cím:Subsite mapping of the binding region of α?amylases with a computer program / Gyöngyi Gyémánt, György Hovánszki, Lili Kandra
Dátum:2002
ISSN:0014-2956 1432-1033
Megjegyzések:A computer program has been evaluated for subsite map calculations of depolymerases. The program runs in WINDOWS and uses the experimentally determined bond cleavage frequencies (BCFs) for determination of the number of subsites, the position of the catalytic site and for calculation of subsite binding energies. The apparent free energy values were optimized by minimization of the differences of the measured and calculated BCF data. The program called SUMA (SUbsite Mapping of alpha-Amylases) is freely available for research and educational purposes via the Internet (E-mail: gyemant@tigris.klte.hu). The advantages of this program are demonstrated through alpha-amylases of different origin, e.g. porcine pancreatic alpha-amylase (PPA) studied in our laboratory, in addition to barley and rice alpha-amylases published in the literature. Results confirm the popular ♭five subsite model' for PPA with three glycone and two aglycone binding sites. Calculations for barley alpha-amylase justify the ♭6 + 2 + (1) model' prediction.The binding area of barleya-amylaseis composed ofsixglycone, twoaglyconebinding sites followedbyabarrier subsite at the reducingendof thebinding site.Calculations for rice alpha-amylase represent an entirely new map with a ♭(1) + 2 + 5 model', where ♭(1)' is a barrier subsite at the nonreducing end of the binding site and there are two glycone and five aglycone binding sites. The rice model may be reminiscent of the action of the bacterial maltogenic amylase, that is, suggesting an exo-mechanism for this enzyme.
Tárgyszavak:Természettudományok Biológiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
subsite mapping
alpha-amylase
action pattern
WINDOWS program
Megjelenés:European Journal of Biochemistry. - 269 : 21 (2002), p. 5157-5162. -
További szerzők:Horánszki György Kandra Lili (1943-) (biokémikus)
Pályázati támogatás:T032005
OTKA
FKFP-0426/2000
Egyéb
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DOI
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2.

001-es BibID:BIBFORM010529
Első szerző:Nielsen, Morten M.
Cím:Two Secondary Carbohydrate Binding Sites on the Surface of Barley alpha-Amylase 1 Have Distinct Functions and Display Synergy in Hydrolysis of Starch Granules / Morten M. Nielsen, Sophie Bozonnet, Eun-Seong Seo, János A. Mótyán, Joakim M. Andersen, Adiphol Dilokpimol, Maher Abou Hachem, Gyöngyi Gyémánt, Henrik Næsted, Lili Kandra, Bent W. Sigurskjold, Birte Svensson
Dátum:2009
ISSN:0006-2960
Megjegyzések:Some polysaccharide processing enzymes possess secondary carbohydrate binding sites situated on the surface far from the active site. In barley alpha-amylase 1 (AMY1), two such sites, SBS1 and SBS2, are found on the catalytic (beta/alpha)8-barrel and the noncatalytic C-terminal domain, respectively. Site-directed mutagenesis of Trp(278) and Trp(279), stacking onto adjacent ligand glucosyl residues at SBS1, and of Tyr(380) and His(395), making numerous ligand contacts at SBS2, suggested that SBS1 and SBS2 act synergistically in degradation of starch granules. While SBS1 makes the major contribution to binding and hydrolysis of starch granules, SBS2 exhibits a higher affinity for the starch mimic beta-cyclodextrin. Compared to that of wild-type AMY1, the K-d of starch granule binding by the SBS1 W278A, W279A, and W278A/W279A mutants thus increased 15-35 times; furthermore, the k(cat)/K-m of W278A/W279A was 2%, whereas both affinity and activity for Y380A at SBS2 were 10% of the wild-type values. Dual site double and triple SBS1/SBS2 substitutions eliminated binding to starch granules, and the k(cat)/K-m of W278A/W279A/Y380A AMY1 was only 0.4% of the wild-type value. Surface plasmon resonance analysis of mutants showed that beta-cyclodextrin binds to SBS2 and SBS1 with K-d,K-1 and K-d,K-2 values of 0.07 and 1.40 mM, respectively. A model that accounts for the observed synergy in starch hydrolysis., where SBS1 and SBS2 bind ordered and free alpha-glucan chains, respectively, thus targeting the enzyme to single alpha-glucan chains accessible for hydrolysis, is proposed. SBS1 and SBS2 also influence the kinetics of hydrolysis for amylose and maltooligosaccharides, the degree of multiple attack on amylose, and subsite binding energies.
Tárgyszavak:Természettudományok Kémiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban
Megjelenés:Biochemistry. - 48 : 32 (2009), p. 7686-7697. -
További szerzők:Bozonnet, Sophie Seo, Eun-Seong Mótyán János András (1981-) (biokémikus, molekuláris biológus) Andersen, Joakim M. Dilokpimol, Adiphol Abou Hachem, Maher Gyémánt Gyöngyi (1960-) (vegyész) Naested, Henrik Kandra Lili (1943-) (biokémikus) Sigurskjold, Bent W. Svensson, Birte
Internet cím:DOI
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3.

001-es BibID:BIBFORM084836
Első szerző:Ramasubbu, Narayanan
Cím:Human salivary alpha-amylase Trp58 situated at subsite -2 is critical for enzyme activity / Narayanan Ramasubbu, Chandran Ragunath, Prasunkumar J. Mishra, Leonard M. Thomas, Gyöngyi Gyémánt, Lili Kandra
Dátum:2004
ISSN:0014-2956 1432-1033
Megjegyzések: The nonreducing end of the substrate?binding site of human salivary α?amylase contains two residues Trp58 and Trp59, which belong to β2-α2 loop of the catalytic (β/α)8 barrel. While Trp59 stacks onto the substrate, the exact role of Trp58 is unknown. To investigate its role in enzyme activity the residue Trp58 was mutated to Ala, Leu or Tyr. Kinetic analysis of the wild?type and mutant enzymes was carried out with starch and oligosaccharides as substrates. All three mutants exhibited a reduction in specific activity (150-180?fold lower than the wild type) with starch as substrate. With oligosaccharides as substrates, a reduction in kcat, an increase in Km and distinct differences in the cleavage pattern were observed for the mutants W58A and W58L compared with the wild type. Glucose was the smallest product generated by these two mutants in the hydrolysis oligosaccharides; in contrast, wild?type enzyme generated maltose as the smallest product. The production of glucose by W58L was confirmed from both reducing and nonreducing ends of CNP?labeled oligosaccharide substrates. The mutant W58L exhibited lower binding affinity at subsites ?2, ?3 and +2 and showed an increase in transglycosylation activity compared with the wild type. The lowered affinity at subsites ?2 and ?3 due to the mutation was also inferred from the electron density at these subsites in the structure of W58A in complex with acarbose?derived pseudooligosaccharide. Collectively, these results suggest that the residue Trp58 plays a critical role in substrate binding and hydrolytic activity of human salivary α?amylase.
Tárgyszavak:Természettudományok Kémiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
Megjelenés:European Journal of Biochemistry. - 271 : 12 (2004), p. 2517-2529. -
További szerzők:Ragunath, Chandran Mishra, Prasunkumar J. Thomas, Leonard M. Gyémánt Gyöngyi (1960-) (vegyész) Kandra Lili (1943-) (biokémikus)
Pályázati támogatás:OTKA T032005
OTKA
OTKA M041829
OTKA
Internet cím:Szerző által megadott URL
DOI
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4.

001-es BibID:BIBFORM081673
Első szerző:Vasas Gábor (biológus-vegyész)
Cím:Capillary Electrophoretic Assay and Purification of Cylindrospermopsin, a Cyanobacterial Toxin from Aphanizomenon ovalisporum, by Plant Test (Blue-Green Sinapis Test) / Gábor Vasas, Attila Gáspár, Gyula Surányi, Gyula Batta, Gyöngyi Gyémánt, Márta M-Hamvas, Csaba Máthé, István Grigorszky, Erika Molnár, George Borbély
Dátum:2002
ISSN:0003-2697
Tárgyszavak:Természettudományok Biológiai tudományok idegen nyelvű folyóiratközlemény külföldi lapban
folyóiratcikk
Megjelenés:Analytical Biochemistry. - 302 : 1 (2002), p. 95-103. -
További szerzők:Gáspár Attila (1970-) (vegyész, kémikus) Surányi Gyula (1957-) (biológus) Batta Gyula (1953-) (molekula-szerkezet kutató) Gyémánt Gyöngyi (1960-) (vegyész) Mikóné Hamvas Márta (1963-) (biológus) Máthé Csaba (1966-) (biológus) Grigorszky István (1967-) (biológus-ökológus) Molnár Erika Borbély György (1943-) (biológus)
Pályázati támogatás:OTKA T5235
OTKA
OTKA T22988
OTKA
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