Purification and characterization of β‐amylase from leaves of potato (Solanum tuberosum)

Abstract

Amylolytic activity is widely distributed in plants. In potato leaves (Solanum tuberosum L.) the abundant amylolytic activity was found to be β‐amylase (EC 3.2.1.2, a‐1,4‐D‐glucan maltohydrolase). β‐Amylase from potato leaves was purified to homogeneity for study of enzyme characteristics. The purification steps included ammonium sulphate precipitation, anion exchange chromatography, affinity chromatography and gel filtration. The end product of α‐1,4‐glucan degradation was maltose. The protein is a 111‐kDa homo‐dimer with a subunit molecular mass of 56 kDa and a pl of 5.6. The pH‐optimum is 6.5 using p‐nitrophenylmaltopentaoside (PNPG5) as substrate. The optimal temperature for hydrolysis is at 40°C. The enzyme is unstable at temperatures above 40°C. The Knt‐value for PNPG5 is 0.73 mM and the activity is inhibited by cyclodextrins. At a concentration of 1 mM, β‐cyclodextrin is a stronger inhibitor than α‐cyclodextrin (68 and 20% inhibition, respectively). Branched glucans (e.g. starch and amylopectin) are superior substrates as compared to long, essentially unbranched glucans (e.g. amylose). This study of the catalytic properties of β‐amylase from potato leaves indicates the importance of β‐amylase as a starch degrading enzyme.

Downregulation of a chloroplast‐targeted β‐amylase leads to a starch‐excess phenotype in leaves

“…Increases in activities of enzymes involved in starch degradation have often been observed in Arabidopsis mutants showing a starch‐excess phenotype (Critchley et al ., 2001; Zeeman et al ., 1998), presumably because the plants attempt to compensate for the decrease in starch degradation. Maltase activity was not detectable in potato leaf extracts, as has been shown previously by Viskø‐Nielsen et al . (1997).…”

Enhancement of umami taste of hydrolyzed protein from wheat gluten by β‐cyclodextrin

“…Previous studies showed that β ‐CD caused variations in activity of many enzymes . However, as seen in Fig.…”

Caracterização parcial de carboidrases, morfologia do grão de amido e composição centesimal de raízes de maca (Lepidium meyenii Walpers)

“…A β-amilase não hidrolisa ligações α-1,6 na amilopectina e sua ação termina no ponto de ramificação; por essa razão, a degradação da amilopectina pela β-amilase é incompleta, formando dextrinas limite de alto peso molecular, que resultam em coloração roxa ao se complexarem com o iodo (Bernfeld, 1951, Whitaker, 1996 A β-amilase está localizada principalmente nos vacúolos de trigo e ervilha (Ziegler, 1999). Vikso et al (1997) (Gana et al, 1998).…”

“…Pires (2005) determinou a atividade enzimática no extrato bruto de mandioquinha de 254 U/mg prot, com aumento da concentração do amido após o terceiro dia pós-colheita e posterior decréscimo, quando armazenados a diferentes temperaturas. Nielsen et al, (1997) encontrou atividades de α-amilase de 16,7U e β-amilase de 54,5U por mg de proteína, numa variedade de batata.…”

“…de saturação, para precipitar β-amilase em lentilha (Lens esculenta L.). Em extratos de folhas de batata (Solanum tuberosum), a maior parte de enzimas amilolíticas precipitou sob concentrações de sulfato de amônio entre 25-50% de saturação (Nielsen et al, 1997). Os valores encontrados foram semelhantes tanto na determinação de atividade total quanto na eletroforese SDS-PAGE, mostrando a homogeneidade entre as frações analisadas.…”