D275P mutant of alcohol dehydrogenase from protozoa Entamoeba histolytica

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D275P mutant of alcohol dehydrogenase from protozoa Entamoeba histolytica

(see also Tetrameric alcohol dehydrogenases and 2nvb)


Publication Abstract from PubMed

Analysis of the three-dimensional structures of two closely related thermophilic and hyperthermophilic alcohol dehydrogenases (ADHs) from the respective microorganisms Entamoeba histolytica (EhADH1) and Thermoanaerobacter brockii (TbADH) suggested that a unique, strategically located proline residue (Pro275) at the center of the dimerization interface might be crucial for maintaining the thermal stability of TbADH. To assess the contribution of Pro275 to the thermal stability of the ADHs, we applied site-directed mutagenesis to replace Asp275 of EhADH1 with Pro (D275P-EhADH1) and conversely Pro275 of TbADH with Asp (P275D-TbADH). The results indicate that replacing Asp275 with Pro significantly enhances the thermal stability of EhADH1 (DeltaT(1/2) </= +10 degrees C), whereas the reverse mutation in the thermophilic TbADH (P275D-TbADH) reduces the thermostability of the enzyme (DeltaT(1/2) </= -18.8 degrees C). Analysis of the crystal structures of the thermostabilized mutant D275P-EhADH1 and the thermocompromised mutant P275D-TbADH suggest that a proline residue at position 275 thermostabilized the enzymes by reducing flexibility and by reinforcing hydrophobic interactions at the dimer-dimer interface of the tetrameric ADHs. Proteins 2008. (c) 2008 Wiley-Liss, Inc.

Thermal stabilization of the protozoan Entamoeba histolytica alcohol dehydrogenase by a single proline substitution., Goihberg E, Dym O, Tel-Or S, Shimon L, Frolow F, Peretz M, Burstein Y, Proteins. 2008 Feb 7;. PMID:18260103

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

Ribbon diagram of the EhADH1 tetramer (2oui). Proline residues (ball representation) are colored orange (Pro275) (which is important for thermal stabilization) and cyan (Pro100). Superposition of the structures of the wild-type apo-EhADH1 (colored lime, 1y9a) and the apo D275P-EhADH1 mutant (colored orange) (2oui). Pro275 and Asp275 are labeled red. Residues within a distance of 4 Å from the mutation are shown (names of monomers are in brackets). Replacing Asp275 with Pro significantly enhanced the thermal stability of EhADH1: ΔT1/260min = +9.3°C, ΔT1/2CD = +10°C. The reverse mutation in the thermophilic TbADH (1ykf; colored magenta) - substitution of wt TbADH Pro275 with Asp (2nvb; colored cyan) reduced the thermal stability of the enzyme: ΔT1/260min = -13.8°C, ΔT1/2CD = -18.8°C. Nitrogen and oxygen atoms are colored in CPK colors. Pro275 and Asp275 are labeled red (names of monomers are in brackets). These findings indicate that a single proline mutation is responsible for the significant differences in the thermal stability of ADHs, and show the importance of prolines in the protein stability. It was also shown that substitution by proline at the important positions could significantly stabilize the protein.


About this Structure

2OUI is a Single protein structure of sequence from Entamoeba histolytica. Full crystallographic information is available from OCA.

Reference

Thermal stabilization of the protozoan Entamoeba histolytica alcohol dehydrogenase by a single proline substitution., Goihberg E, Dym O, Tel-Or S, Shimon L, Frolow F, Peretz M, Burstein Y, Proteins. 2008 Feb 7;. PMID:18260103
Created with the participation of Jaime Prilusky, Michal Harel.

PDB ID 2oui

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Proteopedia Page Contributors and Editors (what is this?)

Alexander Berchansky

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