PDBsum entry 3id4

Hydrolase

PDB id: 3id4

Contents

Protein chain

93 a.a. *

Waters ×70

* Residue conservation analysis

PDB id:

3id4

Name:

Hydrolase

Title:

Crystal structure of rsep pdz2 domain fused gkaspv peptide

Structure:

Regulator of sigma e protease. Chain: a. Fragment: pdz2 domain, residues 222-307. Engineered: yes. Other_details: carboxy-terminally fused gkaspv peptide

Source:

Escherichia coli k-12. Organism_taxid: 83333. Strain: k12. Gene: rsep. Expressed in: escherichia coli. Expression_system_taxid: 469008.

Resolution:

1.60Å R-factor: 0.193 R-free: 0.219

Authors:

X.Li,B.Wang,L.Feng,J.Wang,Y.Shi

Key ref:

X.Li et al. (2009). Cleavage of RseA by RseP requires a carboxyl-terminal hydrophobic amino acid following DegS cleavage. Proc Natl Acad Sci U S A, 106, 14837-14842. PubMed id: 19706448 DOI: 10.1073/pnas.0903289106

Date:

20-Jul-09 Release date: 11-Aug-09

Protein chain

P0AEH1  (RSEP_ECOLI) -  Regulator of sigma-E protease RseP from Escherichia coli (strain K12)

Seq: 450 a.a.

Struc: 93 a.a.*

* PDB and UniProt seqs differ at 6 residue positions (black crosses)

Enzyme reactions

• Enzyme class: E.C.3.4.24.- - ?????

DOI no: 10.1073/pnas.0903289106

Proc Natl Acad Sci U S A 106:14837-14842 (2009)

PubMed id: 19706448

Cleavage of RseA by RseP requires a carboxyl-terminal hydrophobic amino acid following DegS cleavage.

X.Li, B.Wang, L.Feng, H.Kang, Y.Qi, J.Wang, Y.Shi.

ABSTRACT

Regulated intramembrane proteolysis (RIP) by the Site-2 protease (S2P) results in the release of a transmembrane signaling protein. Curiously, however, S2P cleavage must be preceded by the action of the Site-1 protease (S1P). To decipher the underlying mechanism, we reconstituted sequential, in vitro cleavages of the Escherichia coli transmembrane protein RseA by DegS (S1P) and RseP (S2P). After DegS cleavage, the newly exposed carboxyl-terminal residue Val-148 of RseA plays an essential role for RseP cleavage, and its mutation to charged or dissimilar amino acids crippled the Site-2 cleavage. By contrast, the identity of residues 146 and 147 of RseA has no impact on Site-2 cleavage. These results explain why Site-1 cleavage must precede Site-2 cleavage. Structural analysis reveals that the putative peptide-binding groove in the second, but not the first, PDZ domain of RseP is poised for binding to a single hydrophobic amino acid. These observations suggest that after DegS cleavage, the newly exposed carboxyl terminus of RseA may facilitate Site-2 cleavage through direct interaction with the PDZ domain.

Selected figure(s)

Figure 2.
Residue 148 of RseA plays an important role for Site-2 cleavage. (A) Mutation of Val-148 to Thr or Ile, but not His or Lys, allowed Site-2 cleavage. DegS and OMP peptide were added together to the reactions where DegS is indicated. (B) Mutation of Val-148 to Thr or Ile in RseA allowed a more robust envelope stress response than mutation of Val-148 to His or Lys.

Figure 3.
Conserved mutation of Val-148 in RseA allowed retention of Site-2 cleavage. (A) Mutation of Val-148 to conserved, but not dissimilar or charged, amino acids in RseA allowed retention of Site-2 cleavage. DegS and OMP peptide were added together to the reactions where DegS is indicated. (B) Classification of three categories of amino acids at position 148 of RseA based on their impact on Site-1 and Site-2 cleavages. Mutation of Val-148 to any of the five amino acids—Glu, Asp, Gly, Pro, and Phe—crippled Site-1 cleavage of RseA by DegS. Among the mutations that allow Site-1 cleavage, six (mutation of Val-148 to Lys, His, Arg, Ser, Gln, and Tyr) do not allow Site-2 cleavage.

Figures were selected by an automated process.