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| {{STRUCTURE_4cla| PDB=4cla | SCENE= }} | | {{STRUCTURE_4cla| PDB=4cla | SCENE= }} |
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| '''ALTERNATIVE BINDING MODES FOR CHLORAMPHENICOL AND 1-SUBSTITUTED CHLORAMPHENICOL ANALOGUES REVEALED BY SITE-DIRECTED MUTAGENESIS AND X-RAY CRYSTALLOGRAPHY OF CHLORAMPHENICOL ACETYLTRANSFERASE'''
| | ===ALTERNATIVE BINDING MODES FOR CHLORAMPHENICOL AND 1-SUBSTITUTED CHLORAMPHENICOL ANALOGUES REVEALED BY SITE-DIRECTED MUTAGENESIS AND X-RAY CRYSTALLOGRAPHY OF CHLORAMPHENICOL ACETYLTRANSFERASE=== |
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| ==Overview==
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| Leucine-160 of chloramphenicol acetyltransferase (CAT) has been replaced by site-directed mutagenesis to investigate enzyme-ligand interactions at the 1-hydroxyl substituent of the substrate chloramphenicol. The consequences of the substitution of Leu-160 by glutamine and by phenylalanine were deduced from the steady-state kinetic parameters for acetyl transfer from acetyl-CoA to the 3-hydroxyl of chloramphenicol and its analogues 1-deoxychloramphenicol and 1-acetylchloramphenicol. The acetyl group of the latter, which is a substrate both in vivo and in vitro, could potentially bind in a similar position to the 1-hydroxyl of chloramphenicol, in close proximity to the side chain of Leu-160. In the case of Gln-160 CAT, large increases in Km for the three acetyl acceptors were accompanied by small decreases in kcat and in apparent affinity for acetyl-CoA. Such results are consistent with the introduction of the relatively hydrophilic amide in place of the delta-methyl groups of Leu-160. The kinetic properties of Phe-160 CAT were unexpected in that Km for each of the three acetyl acceptors was unchanged or reduced, compared to the equivalent parameters for the wild-type enzyme, whereas kcat fell significantly (44-83-fold) in each case. The ratios of specificity constants (kcat/Km) for the acetylation of chloramphenicol compared with the alternative acyl acceptors were similar for wild-type and mutant enzymes. As the residue substitutions for Leu-160 do not result in enhanced discrimination against the binding and acetylation of 1-acetylchloramphenicol, it appears unlikely that the 1-acetyl group binds to the CAT active site in the same position as that occupied by the 1-hydroxyl of chloramphenicol.(ABSTRACT TRUNCATED AT 250 WORDS)
| | The line below this paragraph, {{ABSTRACT_PUBMED_2015231}}, adds the Publication Abstract to the page |
| | (as it appears on PubMed at http://www.pubmed.gov), where 2015231 is the PubMed ID number. |
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| | {{ABSTRACT_PUBMED_2015231}} |
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| ==About this Structure== | | ==About this Structure== |
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| [[Category: Single protein]] | | [[Category: Single protein]] |
| [[Category: Leslie, A G.W.]] | | [[Category: Leslie, A G.W.]] |
| ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Sun May 4 22:22:10 2008'' | | |
| | ''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Thu Jul 3 13:32:17 2008'' |
