A thermotolerant alkali-resistant gram positive, rod shaped Geobacillus sp.Iso5 was isolated from the thermal water. Identification was performed by biochemical tests and 16S rRNA gene sequencing. The isolate comprises of 55%±0.2 mol % G+C content and iso-C_{15: 0}, iso-C_{16: 0}, iso-C_{17: 0} type of cellular fatty acids. Phylogenetic analysis revealed 98-99% sequence similarities was observed with thermophilic members of group Geobacillus; G.kaustophilus (99.5%), G. thermoleoverance (99%). A hyperthermostable a-amylase was purified using Sephadex G-150 gel filtration chromatography and a DEAE-cellulose column. The SDS-PAGE pattern of purified enzyme was showed an apparent molecular weight of 43 kDa. The MALDI-TOF/TOF mass finger printing analysis of purified enzyme has a 31% close homology with the surface layer protein of Geobacillus sp. The CD and FTIR spectra for secondary structure of enzyme revealed about 55% a –helix, 5% b-strand and 40% of unordered structure. The optimum activity of enzyme was determined to be 90°C and pH 8.0 and can stable up to 90⁰C for 10 min. The DSC-TGA thermogram of purified enzyme was showed only 10% weight loss at 200°C. The effect of EDTA and Zn²⁺ on enzyme activity was shown maximum inhibitory activity. Whereas, Ca²⁺, Mg²⁺, Fe²⁺ and Cu²⁺ were did not affect on the activity of the purified enzyme.