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Mutated barley (1,3...
Mutated barley (1,3)-beta-D-glucan endohydrolases synthesize crystalline (1,3)-beta-D-glucans
- Artikel/kapitelEngelska2002
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Nummerbeteckningar
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LIBRIS-ID:oai:DiVA.org:kth-21831
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https://urn.kb.se/resolve?urn=urn:nbn:se:kth:diva-21831URI
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https://doi.org/10.1074/jbc.M203971200DOI
Kompletterande språkuppgifter
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Språk:engelska
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Sammanfattning på:engelska
Ingår i deldatabas
Klassifikation
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Ämneskategori:ref swepub-contenttype
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Ämneskategori:art swepub-publicationtype
Anmärkningar
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QC 20100525
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Barley (1,3)-beta-D-glucan endohydrolases (EC 3.2.1.39), inactivated by site-directed mutagenesis of their catalytic nucleophiles, show autocondensation glucosynthetic activity with alpha-laminaribiosyl fluoride and heterocondensation glycosynthetic activity with a-laminaribiosyl fluoride and 4'-nitrophenyl beta-D-glucopyranoside. The native enzyme is a retaining endohydrolase of the family 17 group and catalyzes glycosyl transfer reactions at high substrate concentrations. Catalytic efficiencies (k(cat) K-m(-1)) of mutants E231G, E231S, and E231A as glycosynthases are 28.9, 0.9, and 0.5 x 10(-4) M-1 s(-1), respectively. Glycosynthase reactions appear to be processive and proceed with pH optima of 6-8 and yields of up to 75%. Insoluble products formed during the glycosynthase reaction appear as lamellar, hexagonal crystals when observed by electron microscopy. Methylation, NMR, and matrix-assisted laser desorption ionization time-of-flight analyses show that the reaction products are linear (1,3)-beta-D-glucans with a degree of polymerization of 30-34, whereas electron and x-ray diffraction patterns indicate that these (1,3)-beta-D-glucan chains adopt a parallel, triple helical conformation. The (1,3)-beta-D-glucan triple helices are orientated perpendicularly to the plane of the lamellar crystals. The barley (1,3)-beta-D-glucan glycosynthases have considerable potential for tailored and high efficiency synthesis of (1,3)-beta-D-linked oligo- and polysaccharides, some of which could have immunomodulating activity, or for the coupling of (1,3)-beta-D-linked glucosyl residues onto other oligosaccharides or glycoproteins.
Ämnesord och genrebeteckningar
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catalytic amino-acids
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packing analysis
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cdna clone
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in-vitro
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substrate
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polysaccharides
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purification
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(1->3)-beta-d-glucans
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(1->3)-beta-glucan
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glucosidase
Biuppslag (personer, institutioner, konferenser, titlar ...)
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Imai, T.
(författare)
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Rutten, S. J.
(författare)
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Fairweather, J. K.
(författare)
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Pelosi, L.
(författare)
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Bulone, VincentKTH,Bioteknologi(Swepub:kth)u13o1auq
(författare)
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Driguez, H.
(författare)
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Fincher, G. B.
(författare)
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KTHBioteknologi
(creator_code:org_t)
Sammanhörande titlar
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Ingår i:Journal of Biological Chemistry277:33, s. 30102-301110021-92581083-351X
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