PDBsum entry 4mls

Peptide binding protein

PDB id: 4mls

Contents

Protein chains

81 a.a.

12 a.a.

Waters ×61

PDB id:

4mls

Name:

Peptide binding protein

Title:

Crystal structure of the spytag and spycatcher-deltan1 complex

Structure:

Fibronectin binding protein. Chain: a. Fragment: unp residues 461-552. Engineered: yes. Mutation: yes. Spytag. Chain: b. Engineered: yes

Source:

Streptococcus pyogenes. Organism_taxid: 1314. Gene: cnab2, fba2. Expressed in: escherichia coli. Expression_system_taxid: 469008. Synthetic: yes. Other_details: chemically synthesized

Resolution:

1.98Å R-factor: 0.211 R-free: 0.237

Authors:

L.Li,J.O.Fierer,T.A.Rapoport,M.Howarth

Key ref:

L.Li et al. (2014). Structural analysis and optimization of the covalent association between SpyCatcher and a peptide Tag. J Mol Biol, 426, 309-317. PubMed id: 24161952 DOI: 10.1016/j.jmb.2013.10.021

Date:

06-Sep-13 Release date: 13-Nov-13

Protein chain

Q8G9G1  (Q8G9G1_STRPY) -  Fibronectin binding protein from Streptococcus pyogenes

Seq: 733 a.a.

Struc: 81 a.a.*

Protein chain

No UniProt id for this chain

Struc: 12 a.a.

* PDB and UniProt seqs differ at 2 residue positions (black crosses)

DOI no: 10.1016/j.jmb.2013.10.021

J Mol Biol 426:309-317 (2014)

PubMed id: 24161952

Structural analysis and optimization of the covalent association between SpyCatcher and a peptide Tag.

L.Li, J.O.Fierer, T.A.Rapoport, M.Howarth.

ABSTRACT

Peptide tagging is a key strategy for observing and isolating proteins. However, the interactions of proteins with peptides are nearly all rapidly reversible. Proteins tagged with the peptide SpyTag form an irreversible covalent bond to the SpyCatcher protein via a spontaneous isopeptide linkage, thereby offering a genetically encoded way to create peptide interactions that resist force and harsh conditions. Here, we determined the crystal structure of the reconstituted covalent complex of SpyTag and SpyCatcher at 2.1Å resolution. The structure showed the expected reformation of the β-sandwich domain seen in the parental streptococcal adhesin, but flanking sequences at both N- and C-termini of SpyCatcher were disordered. In addition, only 10 out of 13 amino acids of the SpyTag peptide were observed to interact with SpyCatcher, pointing to specific contacts important for rapid split protein reconstitution. Based on these structural insights, we expressed a range of SpyCatcher variants and identified a minimized SpyCatcher, 32 residues shorter, that maintained rapid reaction with SpyTag. Together, these results give insight into split protein β-strand complementation and enhance a distinct approach to ultrastable molecular interaction.