Structural and enzymatic characterization of the lactonase SisLac from Sulfolobus islandicus.
BACKGROUND
A new member of the Phosphotriesterase-Like Lactonases (PLL) family from the hyperthermophilic archeon Sulfolobus islandicus (SisLac) has been characterized. SisLac is a native lactonase that exhibits a high promiscuous phosphotriesterase activity. SisLac thus represents a promising target for engineering studies, exhibiting both detoxification and bacterial quorum quenching abilities, including human pathogens such as Pseudomonas aeruginosa.


RESULTS
Here, we describe the substrate specificity of SisLac, providing extensive kinetic studies performed with various phosphotriesters, esters, N-acyl-homoserine lactones (AHLs) and other lactones as substrates. Moreover, we solved the X-ray structure of SisLac and structural comparisons with the closely related SsoPox structure highlighted differences in the surface salt bridge network and the dimerization interface. SisLac and SsoPox being close homologues (91% sequence identity), we undertook a mutational study to decipher these structural differences and their putative consequences on the stability and the catalytic properties of these proteins.


CONCLUSIONS
We show that SisLac is a very proficient lactonase against aroma lactones and AHLs as substrates. Hence, data herein emphasize the potential role of SisLac as quorum quenching agent in Sulfolobus. Moreover, despite the very high sequence homology with SsoPox, we highlight key epistatic substitutions that influence the enzyme stability and activity.
URMITE UMR CNRS-IRD 6236, IFR48, Faculté de Médecine et de Pharmacie, Université de la Méditerranée, Marseille, France.
Julien
J
Hiblot
Guillaume
G
Gotthard
Eric
E
Chabriere
Mikael
M
Elias
D054742:Acyl-Butyrolactones; D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D004795:Enzyme Stability; D004950:Esterases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D044345:Phosphoric Triester Hydrolases / Q000378:metabolism; D011487:Protein Conformation; D055503:Protein Multimerization; D053038:Quorum Sensing; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D016944:Sulfolobus / Q000201:enzymology* / Q000235:genetics; D013696:Temperature
D054742:Acyl-Butyrolactones; D004950:Esterases; D044345:Phosphoric Triester Hydrolases