PMID- 19646449
OWN - NLM
STAT- MEDLINE
DCOM- 20091020
LR  - 20211020
IS  - 1089-8638 (Electronic)
IS  - 0022-2836 (Print)
IS  - 0022-2836 (Linking)
VI  - 392
IP  - 4
DP  - 2009 Oct 2
TI  - Structure and cleavage specificity of the chymotrypsin-like serine protease 
      (3CLSP/nsp4) of Porcine Reproductive and Respiratory Syndrome Virus (PRRSV).
PG  - 977-93
LID - 10.1016/j.jmb.2009.07.062 [doi]
AB  - Biogenesis and replication of the porcine reproductive and respiratory syndrome 
      virus (PRRSV) include the crucial step of replicative polyprotein processing by 
      self-encoded proteases. Whole genome bioinformatics analysis suggests that 
      nonstructural protein 4 (nsp4) is a 3C-like serine protease (3CLSP), responsible 
      for most of the nonstructural protein processing. The gene encoding this protease 
      was cloned and expressed in Escherichia coli in order to confirm this prediction. 
      The purified protein was crystallized, and the structure was solved at 1.9 A 
      resolution. In addition, the crystal structure of the Ser118Ala mutant was 
      determined at 2.0 A resolution. The monomeric enzyme folds into three domains, 
      similar to that of the homologous protease of equine arteritis virus, which, like 
      PRRSV, is a member of the family Arteriviridae in the order of Nidovirales. The 
      active site of the PRRSV 3CLSP is located between domains I and II and harbors a 
      canonical catalytic triad comprising Ser118, His39, and Asp64. The structure also 
      shows an atypical oxyanion hole and a partially collapsed S1 specificity pocket. 
      The proteolytic activity of the purified protein was assessed in vitro. Three 
      sites joining nonstructural protein domains in the PRRSV replicative polyprotein 
      are confirmed to be processed by the enzyme. Two of them, the nsp3/nsp4 and 
      nsp11/nsp12 junctions, are shown to be cleaved in trans, while cis cleavage is 
      demonstrated for the nsp4/nsp5 linker. Thus, we provide structural evidence as 
      well as enzymatic proof of the nsp4 protein being a functional 3CLSP. We also 
      show that the enzyme has a strong preference for glutamic acid at the P1 position 
      of the substrate.
FAU - Tian, Xinsheng
AU  - Tian X
AD  - CAS Key Laboratory of Pathogenic Microbiology and Immunology, Institute of 
      Microbiology, Chinese Academy of Sciences, Beijing 100101, China.
FAU - Lu, Guangwen
AU  - Lu G
FAU - Gao, Feng
AU  - Gao F
FAU - Peng, Hao
AU  - Peng H
FAU - Feng, Youjun
AU  - Feng Y
FAU - Ma, Guangpeng
AU  - Ma G
FAU - Bartlam, Mark
AU  - Bartlam M
FAU - Tian, Kegong
AU  - Tian K
FAU - Yan, Jinghua
AU  - Yan J
FAU - Hilgenfeld, Rolf
AU  - Hilgenfeld R
FAU - Gao, George F
AU  - Gao GF
LA  - eng
SI  - PDB/3FAN
SI  - PDB/3FAO
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
DEP - 20090729
PL  - Netherlands
TA  - J Mol Biol
JT  - Journal of molecular biology
JID - 2985088R
RN  - 0 (Recombinant Proteins)
RN  - 0 (Viral Proteins)
RN  - EC 3.4.21.- (Serine Endopeptidases)
RN  - EC 3.4.21.1 (Chymotrypsin)
SB  - IM
MH  - Amino Acid Sequence
MH  - Animals
MH  - Catalytic Domain
MH  - Chymotrypsin/chemistry
MH  - Models, Biological
MH  - Models, Molecular
MH  - Molecular Sequence Data
MH  - Porcine respiratory and reproductive syndrome virus/*enzymology
MH  - Protein Structure, Tertiary
MH  - Recombinant Proteins/chemistry/metabolism
MH  - Sequence Homology, Amino Acid
MH  - Serine Endopeptidases/*chemistry/*metabolism
MH  - Structure-Activity Relationship
MH  - Substrate Specificity
MH  - Viral Proteins/chemistry/metabolism
PMC - PMC7094510
EDAT- 2009/08/04 09:00
MHDA- 2009/10/21 06:00
PMCR- 2009/07/29
CRDT- 2009/08/04 09:00
PHST- 2009/05/07 00:00 [received]
PHST- 2009/07/19 00:00 [revised]
PHST- 2009/07/22 00:00 [accepted]
PHST- 2009/08/04 09:00 [entrez]
PHST- 2009/08/04 09:00 [pubmed]
PHST- 2009/10/21 06:00 [medline]
PHST- 2009/07/29 00:00 [pmc-release]
AID - S0022-2836(09)00929-2 [pii]
AID - 10.1016/j.jmb.2009.07.062 [doi]
PST - ppublish
SO  - J Mol Biol. 2009 Oct 2;392(4):977-93. doi: 10.1016/j.jmb.2009.07.062. Epub 2009 
      Jul 29.