PDBsum entry 1qa2

Viral protein/receptor

PDB id: 1qa2

Contents

Protein chain

542 a.a.

Waters ×217

PDB id:

1qa2

Name:

Viral protein/receptor

Title:

Tailspike protein, mutant a334v

Structure:

Tailspike protein. Chain: a. Fragment: receptor binding c-terminal fragment. Engineered: yes. Mutation: yes

Source:

Enterobacteria phage p22. Organism_taxid: 10754

Biol. unit:

Trimer (from PQS)

Resolution:

2.00Å R-factor: not given

Authors:

U.Baxa,S.Steinbacher,A.Weintraub,R.Huber,R.Seckler

Key ref:

U.Baxa et al. (1999). Mutations improving the folding of phage P22 tailspike protein affect its receptor binding activity. J Mol Biol, 293, 693-701. PubMed id: 10543960 DOI: 10.1006/jmbi.1999.3165

Date:

10-Apr-99 Release date: 12-Jan-00

Protein chain

P12528  (FIBER_BPP22) -  Tail spike protein from Salmonella phage P22

Seq: 667 a.a.

Struc: 542 a.a.*

* PDB and UniProt seqs differ at 1 residue position (black cross)

Enzyme reactions

• Enzyme class: E.C.3.2.1.- - ?????

DOI no: 10.1006/jmbi.1999.3165

J Mol Biol 293:693-701 (1999)

PubMed id: 10543960

Mutations improving the folding of phage P22 tailspike protein affect its receptor binding activity.

U.Baxa, S.Steinbacher, A.Weintraub, R.Huber, R.Seckler.

ABSTRACT

Four previously isolated mutations in Salmonella phage P22 tailspike protein were used to study the relationship between protein stability, folding, and function. Tailspike protein binds and hydrolyzes the repetitive O-antigen structure in Salmonella lipopolysaccharide. Four mutations (V331G, V331A, A334V, A334I) are known to increase the folding efficiency, and two of them (at position 331) also increase the thermal stability of the protein. Octasaccharides comprising two repeating units of the O-antigens from two different Salmonella strains were employed to analyze the receptor binding function of the mutant proteins. Their endorhamnosidase enzymatic activity was assayed with the aid of a fluorescence-labeled dodecasaccharide. Both V331A and V331G were found to strongly affect O-antigen binding. Octasaccharide binding affinities of the mutant proteins are reduced tenfold and 200-fold, corresponding to a loss of 17% and 36% of the standard free energy of binding, respectively. Both mutations at position 334 affected O-antigen binding only slightly (DeltaDeltaG(0)B approximately 1 kJ/mol), but these mutations reduce the thermal stability of the protein. The observed effects on the endoglycosidase activity are fully explained by the changes in substrate binding, suggesting that neither of the mutations affect the catalytic rate. Crystal structures of all four mutants were determined to a resolution of 2.0 A. Except for the partly or completely missing side-chain, no significant changes compared to the wild-type protein structure were found for the mutants at position 331, whereas a small but significant backbone displacement around the mutation site in A334V and A334I may explain the observed thermal destabilization.

Selected figure(s)

Figure 1.
Figure 1. Crystal structure of the tailspike protein lacking the head- binding domain complexed with O-antigen octasaccharide. Stereo diagrams of a side view (top) and a cross-section through the b- helices of the trimer (bottom) are depicted. The binding site is shown with octasaccharide only in one subunit, for which only the C a - trace is drawn. The other two subunits are shown in an all atom presentation. Note the solvent- exposed groove in which the saccharide is bound.

Figure 3.
Figure 3. Superpositon of mutant structures with the wt crystal structure in the environment of the mutation site. (a) Structure of A334I with the C a -trace in green and side-chains in magenta, and with the 2jF0 j - jF C j electron den- sity of residues 334 and 335 contoured at 1.5 s level in blue. The superposition of the wt backbone is shown in yel- low with red side-chains. Note the bulging out of the backbone and the difference of Q335 in the mutant structure. The structure of A334V is very similar to the structure of A334I shown here. (b) Superposition of wt (blue), V331A (yellow), and V331G (red). Only residues near the mutation site are shown. The Figures were made using FRODO (Jones et al., 1978) and MOLMOL (Koradi et al., 1996), respectively.

The above figures are reprinted by permission from Elsevier: J Mol Biol (1999, 293, 693-701) copyright 1999.

Figures were selected by an automated process.