Characteristics and thermodynamics of α-amylase from thermophilic actinobacterium, Laceyella sacchari TSI-2. Issue 12 (December 2015)

• Record Type: Journal Article
• Authors: Shukla, R.J.; Singh, S.P.
• Is Part Of: Process biochemistry. Volume 50:Issue 12(2015:Dec.)
• Page Start: 2128
• Page End: 2136
• Publication Date: 2015-12
• Subjects: Laceyella sacchari -- α-Amylase -- Thermodynamic study -- CD spectroscopy -- FT-IR -- HPTLC; Biochemical engineering -- Periodicals; Biotechnology -- Periodicals; Biochemistry -- periodicals; Biotechnology -- periodicals; Chemical Engineering -- periodicals; Génie biochimique -- Périodiques; Biotechnologie -- Périodiques; Biochemical engineering; Biotechnology; Periodicals; 660.63
• DOI: 10.1016/j.procbio.2015.10.013
• Languages: English
• ISSNs: 1359-5113

Abstract

Graphical abstract: Highlights: A highly thermostable α-amylase; half-life-2 h at 100 °C. Amylase stability under extreme conditions of pH, high temperatures and surfactant. Role of bonds and bonding pattern in imparting the thermostability revealed by FT-IR. Changes in content of α-helix and β-sheet probed by CD-spectroscopy suggest effect of temperature on the secondary structure of enzyme. High washing efficiency of the α-amylase. Abstract: A thermophilic actinobacterium, Laceyella sacchari TSI-2 from a hot spring was characterized for its morphological and cultural features. It optimally produced α-amylase at 50 °C and pH 7. The enzyme was purified by 55-fold with 12.23% yield and 4215.91 U/mg specific activity. The molecular weight, K m, V max and K cat were 31 kD, 2.71 mg ml −1, 7.589 μmoles min −1 per mg of the protein and 4.31 × 10 −2 s −1, respectively. Optimum catalysis and stability occurred at 70 °C and pH 7. A stabilizing effect of thiourea was evident, while metal ions and chelators inhibited the enzyme. The enzyme was highly stable in various surfactants. Kd, t 1/2, Δ H *, Δ S * and Δ G * supported thermal stability reflected by the structural integrity and substrate affinity. The HPTLC analysis of the end products suggested the formation of malto-oligosaccharides as intermediate products along with maltose confirming the enzyme as α-amylase. The FT-IR revealed significant role of the triple bonds in thermal stability, while CD spectroscopy suggested changes in the secondary structure of the native and denatured enzymes. The enzyme was highly efficient in removing starch stains from the cotton cloth. Stability at high temperatures, alkaline pH and in surfactants suggests potential of the amylase in various applications.