title: Activation of Bacillus licheniformis alpha-amylase through a disorder-->order transition of the substrate-binding site mediated by a calcium-sodium-calcium metal triad.
abstract: BACKGROUND
  The structural basis as to how metals regulate the functional state of a protein by altering or stabilizing its conformation has been characterized in relatively few cases because the metal-free form of the protein is often partially disordered and unsuitable for crystallographic analysis. This is not the case, however, for Bacillus licheniformis alpha-amylase (BLA) for which the structure of the metal-free form is available. BLA is a hyperthermostable enzyme which is widely used in biotechnology, for example in the breakdown of starch or as a component of detergents. The determination of the structure of BLA in the metal-containing form, together with comparisons to the apo enzyme, will help us to understand the way in which metal ions can regulate enzyme activity.
  
  
  RESULTS
  We report here the crystal structure of native, metal-containing BLA. The structure shows that the calcium-binding site which is conserved in all alpha-amylases forms part of an unprecedented linear triadic metal array, with two calcium ions flanking a central sodium ion. A region around the metal triad comprising 21 residues exhibits a conformational change involving a helix unwinding and a disorder-->order transition compared to the structure of metal-free BLA. Another calcium ion, not previously observed in alpha-amylases, is located at the interface between domains A and C.
  
  
  CONCLUSIONS
  We present a structural description of a major conformational rearrangement mediated by metal ions. The metal induced disorder-->order transition observed in BLA leads to the formation of the extended substrate-binding site and explains on a structural level the calcium dependency of alpha-amylases. Sequence comparisons indicate that the unique Ca-Na-Ca metal triad and the additional calcium ion located between domains A and C might be found exclusively in bacterial alpha-amylases which show increased thermostability. The information presented here may help in the rational design of mutants with enhanced performance in biotechnological applications.
authors.affiliation: Max-Planck-Institut für Biochemie, Planegg-Martinsried, Germany. machius@chop.swmed.edu
authors.forename: M
authors.initials: M
authors.lastname: Machius
authors.forename: N
authors.initials: N
authors.lastname: Declerck
authors.forename: R
authors.initials: R
authors.lastname: Huber
authors.forename: G
authors.initials: G
authors.lastname: Wiegand
mesh_terms: D001407:Bacillus / Q000201:enzymology*; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry / Q000378:metabolism*; D018360:Crystallography, X-Ray; D004795:Enzyme Stability; D008958:Models, Molecular; D011487:Protein Conformation; D017510:Protein Folding; D012964:Sodium / Q000737:chemistry / Q000378:metabolism*; D000516:alpha-Amylases / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D012964:Sodium; D000516:alpha-Amylases; D002118:Calcium