@article {65022,
	title = {Large scale structural rearrangement of a serine hydrolase from Francisella tularensis facilitates catalysis.},
	journal = {J Biol Chem},
	volume = {288},
	year = {2013},
	month = {2013 Apr 12},
	pages = {10522-35},
	abstract = {<p>Tularemia is a deadly, febrile disease caused by infection by the gram-negative bacterium, Francisella tularensis. Members of the ubiquitous serine hydrolase protein family are among current targets to treat diverse bacterial infections. Herein we present a structural and functional study of a novel bacterial carboxylesterase (FTT258) from F. tularensis, a homologue of human acyl protein thioesterase (hAPT1). The structure of FTT258 has been determined in multiple forms, and unexpectedly large conformational changes of a peripheral flexible loop occur in the presence of a mechanistic cyclobutanone ligand. The concomitant changes in this hydrophobic loop and the newly exposed hydrophobic substrate binding pocket suggest that the observed structural changes are essential to the biological function and catalytic activity of FTT258. Using diverse substrate libraries, site-directed mutagenesis, and liposome binding assays, we determined the importance of these structural changes to the catalytic activity and membrane binding activity of FTT258. Residues within the newly exposed hydrophobic binding pocket and within the peripheral flexible loop proved essential to the hydrolytic activity of FTT258, indicating that structural rearrangement is required for catalytic activity. Both FTT258 and hAPT1 also showed significant association with liposomes designed to mimic bacterial or human membranes, respectively, even though similar structural rearrangements for hAPT1 have not been reported. The necessity for acyl protein thioesterases to have maximal catalytic activity near the membrane surface suggests that these conformational changes in the protein may dually regulate catalytic activity and membrane association in bacterial and human homologues.</p>
},
	keywords = {Bacterial Proteins, Cell Membrane, Francisella tularensis, Humans, Mutagenesis, Site-Directed, Serine Endopeptidases, Structural Homology, Protein, Structure-Activity Relationship, Thiolester Hydrolases},
	issn = {1083-351X},
	doi = {10.1074/jbc.M112.446625},
	author = {Filippova, Ekaterina V and Weston, Leigh A and Kuhn, Misty L and Geissler, Brett and Gehring, Alexandra M and Armoush, Nicola and Adkins, Chinessa T and Minasov, George and Dubrovska, Ievgeniia and Shuvalova, Ludmilla and Winsor, James R and Lavis, Luke D and Satchell, Karla J F and Becker, Daniel P and Anderson, Wayne F and Johnson, R Jeremy}
}
@article {6299,
	title = {The structural basis for the narrow substrate specificity of an acetyl esterase from Thermotoga maritima.},
	journal = {Biochimica et Biophysica Acta},
	volume = {1824},
	year = {2012},
	month = {2012 Sep},
	pages = {1024-30},
	abstract = {<p>Acetyl esterases from carbohydrate esterase family 7 exhibit unusual substrate specificity. These proteins catalyze the cleavage of disparate acetate esters with high efficiency, but are unreactive to larger acyl groups. The structural basis for this distinct selectivity profile is unknown. Here, we investigate a thermostable acetyl esterase (TM0077) from Thermotoga maritima using evolutionary relationships, structural information, fluorescent kinetic measurements, and site directed mutagenesis. We measured the kinetic and structural determinants for this specificity using a diverse series of small molecule enzyme substrates, including novel fluorogenic esters. These experiments identified two hydrophobic plasticity residues (Pro228, and Ile276) surrounding the nucleophilic serine that impart this specificity of TM0077 for small, straight-chain esters. Substitution of these residues with alanine imparts broader specificity to TM0077 for the hydrolysis of longer and bulkier esters. Our results suggest the specificity of acetyl esterases have been finely tuned by evolution to catalyze the removal of acetate groups from diverse substrates, but can be modified by focused amino acid substitutions to yield enzymes capable of cleaving larger ester functionalities.</p>
},
	keywords = {Acetylesterase, Bacterial Proteins, Binding Sites, Catalysis, Esters, Fluoresceins, Fluorescent Dyes, Hydrolysis, Kinetics, Models, Molecular, Mutagenesis, Site-Directed, Substrate Specificity, Thermotoga maritima},
	issn = {0006-3002},
	doi = {10.1016/j.bbapap.2012.05.009},
	author = {Hedge, Matthew K and Gehring, Alexandra M and Adkins, Chinessa T and Weston, Leigh A and Lavis, Luke D and Johnson, R Jeremy}
}
