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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
Internet cím:	Szerző által megadott URL DOI Intézményi repozitóriumban (DEA) tárolt változat
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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 Intézményi repozitóriumban (DEA) tárolt változat
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