AbstractAp‐nitrophenyl‐α‐D‐glycopyranoside‐hydrolyzing oligo‐1,6‐glucosidase (dextrin 6‐α‐glucanohydrolase, EC 3.2.1.10) of a caldoactiveBacillussp. KP 1228 capable of growing at 51–82°C was purified to homogeneity. The molecular weight was estimated as 140,000. The enzyme consisted of two identical subunits each comprising a threonine residue at the NH2‐terminus. The enzyme was most active at 85°C and pH 5.1, and stable for 10 min up to 85°C at pH 6.8. The enzyme had no antigenic determinant common to oligo‐1,6‐glucosidases fromBacillus cereusATCC 7064 (mesophile),Bacillus coagulansATCC 7050 (facultative thermophile) andBacillus thermoglucosidasiusKP 1006 (DSM 2542) (obligate thermophile). A strong correlation between the increase in proline content and the rise in thermostability of