Crystallographic substrate binding studies of Leishmania mexicana SCP2-thiolase (type-2): unique features of oxyanion hole-1

Abstract Structures of the C123A variant of the dimeric Leishmania mexicana SCP2-thiolase (type-2) (Lm-thiolase), complexed with acetyl-CoA and acetoacetyl-CoA, respectively, are reported. The catalytic site of thiolase contains two oxyanion holes, OAH1 and OAH2, which are important for catalysis. T...

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Bibliographic Details
Published in:Protein engineering, design and selection design and selection, 2017-03, Vol.30 (3), p.227-236
Main Authors: Harijan, Rajesh K., Kiema, Tiila-Riikka, Syed, Shahan M., Qadir, Imran, Mazet, Muriel, Bringaud, Frédéric, Michels, Paul A.M., Wierenga, Rik K.
Format: Article
Language:English
Subjects:
Acetyl-CoA C-Acetyltransferase - chemistry
Biochemistry, Molecular Biology
Catalytic Domain
Crystallography, X-Ray
Leishmania mexicana - enzymology
Life Sciences
Microbiology and Parasitology
Parasitology
Protein Structure, Secondary
Protozoan Proteins - chemistry
Structural Biology
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Summary:Abstract Structures of the C123A variant of the dimeric Leishmania mexicana SCP2-thiolase (type-2) (Lm-thiolase), complexed with acetyl-CoA and acetoacetyl-CoA, respectively, are reported. The catalytic site of thiolase contains two oxyanion holes, OAH1 and OAH2, which are important for catalysis. The two structures reveal for the first time the hydrogen bond interactions of the CoA-thioester oxygen atom of the substrate with the hydrogen bond donors of OAH1 of a CHH-thiolase. The amino acid sequence fingerprints (CxS, NEAF, GHP) of three catalytic loops identify the active site geometry of the well-studied CNH-thiolases, whereas SCP2-thiolases (type-1, type-2) are classified as CHH-thiolases, having as corresponding fingerprints CxS, HDCF and GHP. In all thiolases, OAH2 is formed by the main chain NH groups of two catalytic loops. In the well-studied CNH-thiolases, OAH1 is formed by a water (of the Wat-Asn(NEAF) dyad) and NE2 (of the GHP-histidine). In the two described liganded Lm-thiolase structures, it is seen that in this CHH-thiolase, OAH1 is formed by NE2 of His338 (HDCF) and His388 (GHP). Analysis of the OAH1 hydrogen bond networks suggests that the GHP-histidine is doubly protonated and positively charged in these complexes, whereas the HDCF histidine is neutral and singly protonated.
ISSN:1741-0126
1741-0134
DOI:10.1093/protein/gzw080