title: Structural and functional characterization of an anesthetic binding site in the second cysteine-rich domain of protein kinase Cδ*.
abstract: Elucidating the principles governing anesthetic-protein interactions requires structural determinations at high resolutions not yet achieved with ion channels. Protein kinase C (PKC) activity is modulated by general anesthetics. We solved the structure of the phorbol-binding domain (C1B) of PKCδ complexed with an ether (methoxymethylcycloprane) and with an alcohol (cyclopropylmethanol) at 1.36-Å resolution. The cyclopropane rings of both agents displace a single water molecule in a surface pocket adjacent to the phorbol-binding site, making van der Waals contacts with the backbone and/or side chains of residues Asn-237 to Ser-240. Surprisingly, two water molecules anchored in a hydrogen-bonded chain between Thr-242 and Lys-260 impart elasticity to one side of the binding pocket. The cyclopropane ring takes part in π-acceptor hydrogen bonds with the amide of Met-239. There is a crucial hydrogen bond between the oxygen atoms of the anesthetics and the hydroxyl of Tyr-236. A Tyr-236-Phe mutation results in loss of binding. Thus, both van der Waals interactions and hydrogen-bonding are essential for binding to occur. Ethanol failed to bind because it is too short to benefit from both interactions. Cyclopropylmethanol inhibited phorbol-ester-induced PKCδ activity, but failed to do so in PKCδ containing the Tyr-236-Phe mutation.
authors.affiliation: Department of Anesthesia and Critical Care, Massachusetts General Hospital, Boston, MA, USA.
authors.forename: Sivananthaperumal
authors.initials: S
authors.lastname: Shanmugasundararaj
authors.forename: Joydip
authors.initials: J
authors.lastname: Das
authors.forename: Warren S
authors.initials: WS
authors.lastname: Sandberg
authors.forename: Xiaojuan
authors.initials: X
authors.lastname: Zhou
authors.forename: Dan
authors.initials: D
authors.lastname: Wang
authors.forename: Robert O
authors.initials: RO
authors.lastname: Messing
authors.forename: Karol S
authors.initials: KS
authors.lastname: Bruzik
authors.forename: Thilo
authors.initials: T
authors.lastname: Stehle
authors.forename: Keith W
authors.initials: KW
authors.lastname: Miller
mesh_terms: D018681:Anesthetics, General / Q000737:chemistry*; D001665:Binding Sites; D003521:Cyclopropanes / Q000737:chemistry*; D003545:Cysteine / Q000737:chemistry*; D004987:Ethers / Q000737:chemistry*; D006801:Humans; D000432:Methanol / Q000031:analogs & derivatives* / Q000737:chemistry; D011485:Protein Binding; D011487:Protein Conformation; D051745:Protein Kinase C-delta / Q000737:chemistry* / Q000648:ultrastructure*; D017434:Protein Structure, Tertiary; D013329:Structure-Activity Relationship
chemical_list: D018681:Anesthetics, General; D003521:Cyclopropanes; D004987:Ethers; C490695:cyclopropylcarbinol; D051745:Protein Kinase C-delta; D003545:Cysteine; D000432:Methanol