The complete nucleotide sequence encoding the high-molecular-mass amylase (HMMA) ofGeobacillus stearothermophilusATCC 12980 was established by PCR techniques. Based on thehmmagene sequence, the full-length rHMMA, four N- or C-terminal rHMMA truncations as well as three C-terminal rHMMA fragments were cloned and heterologously expressed inEscherichia coli.Purified rHMMA forms were used either for affinity studies with the recombinant (r) S-layer protein SbsC (rSbsC), peptidoglycan-containing sacculi (PGS) and pure peptidoglycan (PG) devoid of the secondary cell wall polymer (SCWP), or for surface plasmon resonance (SPR) studies using rSbsC and isolated SCWP. In the C-terminal part of the HMMA, three specific binding regions, one for each cell wall component (rSbsC, SCWP and PG), could be identified. The functionality of the PG-binding domain could be confirmed by replacing the main part of the SCWP-binding domain of an S-layer protein by the PG-binding domain of the HMMA. The present work describes a completely new and highly economic strategy for cell adhesion of an exoenzyme.