A new serine protease fold revealed by the crystal structure of human cytomegalovirus protease

Abstract

A new polypeptide backbone fold for serine proteases has been identified based on the crystal structure ofhmnan cytomcgalovi.J.us protease. The structure was detem1ined at 2.5A resolution by the multiplewavelength anomalous diJ'fraction technique using the seleno-methionyl protein and refined at 2.0A resolution. It reveals a seven-stnmded mostly-antipm·allel 13-ban·eL which is sunounded by seven helices. The active site residues (Ser-132 and His-63) m·e situated on the outside of the 13-bmrel and in a groove on the surface of the protein. TI1e stmctme suggests that the third member of the catalytic triad is probably His-157. The protease of herpesviruses plays an essential role in the production of lllfectious virions 311d is an attractive tmget for the developement of antiherpes agents. The crystal stmcture information vvill help in the design m1d optimization of inhibitors agai.J.1st herpes virus protease.[1] Tong, L., Qian, C., Massariol, M. Human cytomegalovirus (HCMV) is a beta herpes vi.J.us. HCJ\.1Y, like all other members of the Herpes virus family, encodes a protease that is essential for capsid maturation and production of infectious virus. TI1e catalytic domain of the HCMV protease was produced in E.coli as a single-chain protein and was crystallized in space group C222 1 vvith two dimers per asymmetric unit. TI1e crystal stmcrure was deteffili.ned at 2.5A resolution using the ISAS method m1d noncryst:1llographico symmetry averaging. Om cmTent model has been refined against 2.3A data collected on an in1age plate at -170°C. The HCMV protease structure has a new fold different from that of any other 1.!1own protease. There is a central core comprising two orthogonal4-stranded beta sheets sunounded by eight alpha-helices. Residues in three flexible surface loops, including two associated with internal cleavage sites at ill11ino acids 143 and 209, have not been modeled into the wnent stmcture. Dimerization ofHCMV protease is mediated primarily by bmying four turns of alphaheli" (218-232) from one monomer into a pronounced depression in the smface of the other monomer. Only two of three residues previously in1plicated by ill11ino acid sequence alignment and mutagenesis asparticipating directly in catalysis (Ser-132 and His-63 but not Glu-122) me actually located in the active site. This novel structure is being used to finther understand the catalytic mechllilism. and to design inhibitors as potential anti-viral agents. Cysteine proteases related to mmnmalian inter!eukin-1~ convertlilg enzyme (ICE) and to its C. elegans homologue, CED-3, play a critical role in the biochemical events tl1at culminate in apoptosis. We have deteffili.ned the three-dimensional stmctme of a complex of tl1e hmnan CED-3 homologue CPP32/apopain witll a potent tetrapeptide-aldehyde inhibitor. I TI1e protein resembles ICE in overall structure, but its S 4 subsite is strikingly different in size and chemical composition. These differences account for the vmiation in specificity between tl1e ICE-and CED-3-related proteases and enable ...

Unconventional serine proteases: Variations on the catalytic Ser/His/Asp triad configuration

“…The Ser/His/His catalytic triad was validated by the crystal structure of the cytomegalovirus protease (Qiu et al 1996;Shieh et al 1996;Tong et al 1996). The structure revealed that the protein has a unique fold with the Ser132 Og atom and bridging His63 Ne2 nitrogen being within hydrogen-bonding distance (Fig.…”

Complex of NS3 protease and NS4A peptide of BK strain hepatitis C virus: A 2.2 Å resolution structure in a hexagonal crystal form

“…Each subunit of this protease contains a catalytic triad that maintains a similar geometry as seen for other serine proteases. On the other hand, each subunit is a single domain a/P protein and processes a different overall fold from other members of the serine protease family (Chen et al, 1996;Qiu et al, 1996;Shieh et al, 1996;Tong et al, 1996).…”

Assemblins as maturational proteases in herpesviruses

“…2) [58,59]. The stabilization of the oxyanion hole is realized in a similar manner in all serine proteases as well.…”

“…The first crystal structures of an assemblin were published by three groups in 1996 [58,59,83]. All three structures are of dimeric HCMV-assemblin.…”