A thermophilic actinobacterium, Laceyella sacchari TSI-2 from a hot spring was characterized for its morphological and cultural features. It optimally produced alpha-amylase at 50 degrees 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 mu moles min(-1) per mg of the protein and 4.31 x 10(-2) s(-1), respectively. Optimum catalysis and stability occurred at 70 degrees 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), Delta H*, Delta S* and Delta G* supported thermal stability reflected by the structural integrity and substrate affinity. The HPTLC analysis of the end products suggested the formation of maltooligosaccharides as intermediate products along with maltose confirming the enzyme as alpha-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. (C) 2015 Elsevier Ltd. All rights reserved.