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Titolo:
Mutations of barley beta-amylase that improve substrate-binding affinity and thermostability
Autore:
Ma, YF; Evans, DE; Logue, SJ; Langridge, P;
Indirizzi:
Univ Adelaide, Dept Plant Sci, Glen Osmond, SA 5064, Australia Univ Adelaide Glen Osmond SA Australia 5064 en Osmond, SA 5064, Australia IMVS, Hanson Ctr Canc Res, Div Human Immunol, Adelaide, SA 5000, AustraliaIMVS Adelaide SA Australia 5000 man Immunol, Adelaide, SA 5000, Australia
Titolo Testata:
MOLECULAR GENETICS AND GENOMICS
fascicolo: 3, volume: 266, anno: 2001,
pagine: 345 - 352
SICI:
1617-4615(200111)266:3<345:MOBBTI>2.0.ZU;2-P
Fonte:
ISI
Lingua:
ENG
Soggetto:
HORDEUM-VULGARE L.; LIMIT DEXTRINASE; SEVENFOLD-MUTANT; SWISS-MODEL; ACTIVE-SITE; EXPRESSION; RESOLUTION; VARIETIES; MALTOSE;
Keywords:
mutation; beta-amylase; barley; thermostability; substrate-binding affinity;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Life Sciences
Citazioni:
26
Recensione:
Indirizzi per estratti:
Indirizzo: Ma, YF Univ Adelaide, Dept Plant Sci, Waite Campus, Glen Osmond, SA 5064, Australia Univ Adelaide Waite Campus Glen Osmond SA Australia 5064 Australia
Citazione:
Y.F. Ma et al., "Mutations of barley beta-amylase that improve substrate-binding affinity and thermostability", MOL GENET G, 266(3), 2001, pp. 345-352

Abstract

Three allelic forms of barley beta -amylase (Sd1, Sd2H and Sd2L) exhibit different thermostability and kinetic properties. These differences critically influence the malting quality of barley varieties. To understand the molecular basis for the different properties of these three allelic forms, Sd1and Sd2L beta -amylase cDNAs were cloned, and the effects of the amino acid substitutions between them were evaluated by site-directed mutagenesis. The results showed that an R115C mutation is responsible for the difference in kinetic properties. This substitution resulted in an additional hydrogenbond which may create a more favourable environment for substrate-binding. The different thermostabilities of the beta -amylase forms are due to two amino acid substitutions (V233A and L347S), which increased the enzyme's thermostability index T-50 by 1.9 degreesC and 2.1 degreesC, respectively. The increased thermostability associated with these two mutations may be due to relief of steric strain and the interaction of the protein surface with solvent water. Although both V233A and L347S mutations increased thermostability, they affected the thermostability in different ways. The replacementof L347 by serine seems to increase the thermostability by slowing thermalunfolding of the protein during heating, while the replacement of V233 by alanine appears to cause an acceleration of the refolding after heating. Because the different beta -amylase properties determined by the three mutations (R115C, V233A and L347S) are associated with malting quality of barley variety, a mutant with high thermostability and substrate-binding affinity was generated by combining the three preferred amino acid residues C115, A233 and S347 together. A possible approach to producing barley varieties with better malting quality by genetic engineering is discussed.

ASDD Area Sistemi Dipartimentali e Documentali, Università di Bologna, Catalogo delle riviste ed altri periodici
Documento generato il 04/04/20 alle ore 12:01:29