In order to understand the role of N1 domain (1-257 aa) in the amylopullulanase (gt-apu) of the extremely thermophilic bacterium Geobacillus thermoleovorans NP33, N1 deletion construct (gt-apu Delta N) has been generated and expressed in Escherichia coli. The truncated amylopullulanase (gt-apu Delta N) exhibits similar pH and temperature optima like gt-apu, but enhanced thermostability. The gt-apu Delta N has greater hydrolytic action and specific activity on pullulan than gt-apu. The k(cat) (starch and pullulan) and K-m (starch) values of gt-apu Delta N increased, while K-m (pullulan) decreased. The enzyme upon N1 deletion hydrolyzed maltotetraose as the smallest substrate in contrast to maltopentaose of gt-apu. The role of N1 domain of gt-apu in raw starch binding has been confirmed, for the first time, based on deletion and Langmuir-Hinshelwood kinetics. Furthermore, N1 domain appears to exert a negative influence on the thermostability of gt-apu because N1 truncation significantly improves thermostability.