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| - | [[Image:1a40.gif|left|200px]] | + | {{Seed}} |
| | + | [[Image:1a40.png|left|200px]] |
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| | {{STRUCTURE_1a40| PDB=1a40 | SCENE= }} | | {{STRUCTURE_1a40| PDB=1a40 | SCENE= }} |
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| - | '''PHOSPHATE-BINDING PROTEIN WITH ALA 197 REPLACED WITH TRP'''
| + | ===PHOSPHATE-BINDING PROTEIN WITH ALA 197 REPLACED WITH TRP=== |
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| - | ==Overview==
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| - | Stringent specificity and complementarity between the receptor, a periplasmic phosphate-binding protein (PBP) with a two-domain structure, and the completely buried and dehydrated phosphate are achieved by hydrogen bonding or dipolar interactions. We recently found that the surface charge potential of the cleft between the two domains that contains the anion binding site is intensely electronegative. This novel finding prompted the study reported here of the effect of ionic strength on the equilibrium and rapid kinetics of phosphate binding. To facilitate this study, Ala197, located on the edge of the cleft, was replaced by a Trp residue (A197W PBP) to generate a fluorescence reporter group. The A197W PBP-phosphate complex retains wild-type Kd and X-ray structure beyond the replacement residue. The Kd (0.18 microM) at no salt is increased by 20-fold at greater than 0.30 M NaCl. Stopped-flow fluorescence kinetic studies indicate a two-step binding process: (1) The phosphate (L) binds, at near diffusion-controlled rate, to the open cleft form (Po) of PBP to produce an intermediate, PoL. This rate decreases with increasing ionic strength. (2) The intermediate isomerizes to the closed-conformation form, PcL. The results indicate that the high specificity, affinity, and rate of phosphate binding are not influenced by the noncomplementary electronegative surface potential of the cleft. That binding depends almost entirely on local dipolar interactions with the receptor has important ramification in electrostatic interactions in protein structures and in ligand recognition.
| + | The line below this paragraph, {{ABSTRACT_PUBMED_9865949}}, adds the Publication Abstract to the page |
| | + | (as it appears on PubMed at http://www.pubmed.gov), where 9865949 is the PubMed ID number. |
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| | + | {{ABSTRACT_PUBMED_9865949}} |
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| | ==About this Structure== | | ==About this Structure== |
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| | [[Category: Mutagenesis]] | | [[Category: Mutagenesis]] |
| | [[Category: Phosphate-binding protein]] | | [[Category: Phosphate-binding protein]] |
| - | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Fri May 2 09:46:43 2008'' | + | |
| | + | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Mon Jun 30 15:58:58 2008'' |
Revision as of 12:59, 30 June 2008
Template:STRUCTURE 1a40
PHOSPHATE-BINDING PROTEIN WITH ALA 197 REPLACED WITH TRP
Template:ABSTRACT PUBMED 9865949
About this Structure
1A40 is a Single protein structure of sequence from Escherichia coli. Full crystallographic information is available from OCA.
Reference
Dominant role of local dipolar interactions in phosphate binding to a receptor cleft with an electronegative charge surface: equilibrium, kinetic, and crystallographic studies., Ledvina PS, Tsai AL, Wang Z, Koehl E, Quiocho FA, Protein Sci. 1998 Dec;7(12):2550-9. PMID:9865949
Page seeded by OCA on Mon Jun 30 15:58:58 2008
