PDBsum entry 1jd9

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Hydrolase PDB id
Protein chain
448 a.a. *
Waters ×190
* Residue conservation analysis

References listed in PDB file
Key reference
Title Structural basis of alpha-Amylase activation by chloride.
Authors N.Aghajari, G.Feller, C.Gerday, R.Haser.
Ref. Protein Sci, 2002, 11, 1435-1441. [DOI no: 10.1110/ps.0202602]
PubMed id 12021442
To further investigate the mechanism and function of allosteric activation by chloride in some alpha-amylases, the structure of the bacterial alpha-amylase from the psychrophilic micro-organism Pseudoalteromonas haloplanktis in complex with nitrate has been solved at 2.1 A degrees, as well as the structure of the mutants Lys300Gln (2.5 A degrees ) and Lys300Arg (2.25 A degrees ). Nitrate binds strongly to alpha-amylase but is a weak activator. Mutation of the critical chloride ligand Lys300 into Gln results in a chloride-independent enzyme, whereas the mutation into Arg mimics the binding site as is found in animal alpha-amylases with, however, a lower affinity for chloride. These structures reveal that the triangular conformation of the chloride ligands and the nearly equatorial coordination allow the perfect accommodation of planar trigonal monovalent anions such as NO3-, explaining their unusual strong binding. It is also shown that a localized negative charge such as that of Cl-, rather than a delocalized charge as in the case of nitrate, is essential for maximal activation. The chloride-free mutant Lys300Gln indicates that chloride is not mandatory for the catalytic mechanism but strongly increases the reactivity at the active site. Disappearance of the putative catalytic water molecule in this weakly active mutant supports the view that chloride helps to polarize the hydrolytic water molecule and enhances the rate of the second step in the catalytic reaction.
Figure 1.
Fig. 1. Schematic representation of the chloride binding site and of the interaction network with active site residues (adapted from Qian et al. 1994). The essential catalytic residues are in italics. Chloride ligands are Lys300(337), Arg172(195), Asn262(298), and H[2]O1003(525); active site residues are Glu200(233), Asp174(197), and Asp264(300) the putative catalytic water molecule H[2]O1004(cat) and His263(299). Numbers in parentheses refer to pig pancreas -amylase.
Figure 2.
Fig. 2. Superposition of the chloride binding site in P. haloplanktis -amylase (AHA), in green, complex AHA/nitrate in blue, mutant K300Q in yellow, and mutant K300R in pink.
The above figures are reprinted by permission from the Protein Society: Protein Sci (2002, 11, 1435-1441) copyright 2002.
Secondary reference #1
Title Structures of the psychrophilic alteromonas haloplanctis alpha-Amylase give insights into cold adaptation at a molecular level.
Authors N.Aghajari, G.Feller, C.Gerday, R.Haser.
Ref. Structure, 1998, 6, 1503-1516. [DOI no: 10.1016/S0969-2126(98)00149-X]
PubMed id 9862804
Full text Abstract
Figure 4.
Figure 4. A representation of charges at the surfaces of (a) AHA, (b) HPA and (c) BLA, displayed at the same potential range. Color codes are: red, aspartic and glutamic acids; blue, lysines and arginines. This figure was generated with the program GRASP [57].
The above figure is reproduced from the cited reference with permission from Cell Press
Secondary reference #2
Title Crystal structures of the psychrophilic alpha-Amylase from alteromonas haloplanctis in its native form and complexed with an inhibitor.
Authors N.Aghajari, G.Feller, C.Gerday, R.Haser.
Ref. Protein Sci, 1998, 7, 564-572. [DOI no: 10.1002/pro.5560070304]
PubMed id 9541387
Full text Abstract
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