The yeast Lipomyces kononenkoae (Lk) secretes a highly active raw starch-degrading α-amylase (αAmy) that liberates reducing groups from glucose polymers containing both α-1,4 and α-1,6 bonds. The LKA1 gene encoding this industrially important αAmy was cloned as a 2261-bp cDNA fragment from a glucose-derepressed mutant (IGC4052B) of Lk and characterized. The nucleotide (nt) sequence of the cDNA fragment was determined, revealing an open reading frame of 1872 bp, encoding a 596 amino-acid (aa) mature protein (LKA1) with a calculated Mr of 65706. The similarity between the aa sequence of LKA1 and those of other αAmy showed four common conserved regions characteristic of the αAmy protein family: (A) 264DIVVNH269, (B) 349GLRIDTVKH357, (B') 376GEVFD380 and (C) 439FLENQD444. The deduced aa sequence revealed significant homology to the aa sequences of the Aspergillus oryzae, Schwanniomyces occidentalis and Saccharomycopsis fibuligera αAmy, various bacterial cyclomaltodextrin glucanotransferases, a β-amylase and the 5′-region of a glucoamylase. LKA1 was expressed in Saccharomyces cerevisiae (Sc) under the control of the phosphoglycerate kinase (PGK1) promoter and Northern blot analysis showed the presence of a single 2.3-kb transcript. The 28-aa signal peptide of the LKA1 protein efficiently directed its secretion into the medium when expressed in Sc.