Distant Homology Modeling of LCAT and Its Validation through In Silico Targeting and In Vitro and In Vivo Assays

作者：Sensi Cristina; Simonelli Sara; Zanotti Ilaria; Tedeschi Gabriella; Lusardi Giulia; Franceschini Guido; Calabresi Laura; Eberini Ivano^*

来源：PLos One, 2014, 9(4): e95044.

DOI：10.1371/journal.pone.0095044

LCAT (lecithin: cholesterol acyltransferase) catalyzes the transacylation of a fatty acid of lecithin to cholesterol, generating a cholesteryl ester and lysolecithin. The knowledge of LCAT atomic structure and the identification of the amino acids relevant in controlling its structure and function are expected to be very helpful to understand the enzyme catalytic mechanism, as involved in HDL cholesterol metabolism. However - after an early report in the late %26apos;90 s - no recent advance has been made about LCAT three-dimensional structure. In this paper, we propose an LCAT atomistic model, built following the most up-to-date molecular modeling approaches, and exploiting newly solved crystallographic structures. LCAT shows the typical folding of the alpha/beta hydrolase superfamily, and its topology is characterized by a combination of alpha-helices covering a central 7-strand beta-sheet. LCAT presents a Ser/Asp/His catalytic triad with a peculiar geometry, which is shared with such other enzyme classes as lipases, proteases and esterases. Our proposed model was validated through different approaches. We evaluated the impact on LCAT structure of some point mutations close to the enzyme active site (Lys218Asn, Thr274Ala, Thr274Ile) and explained, at a molecular level, their phenotypic effects. Furthermore, we devised some LCAT modulators either designed through a de novo strategy or identified through a virtual high-throughput screening pipeline. The tested compounds were proven to be potent inhibitors of the enzyme activity.

• 出版日期2014-4-15