PDBsum entry 2rtj

Biotin-binding protein

PDB id: 2rtj

Contents

Protein chain

123 a.a. *

Ligands

GLL

FMT

Waters ×132

* Residue conservation analysis

PDB id:

2rtj

Name:

Biotin-binding protein

Title:

Streptavidin-glycoluril complex, ph 2.50, space group i4122

Structure:

Streptavidin. Chain: a

Source:

Streptomyces avidinii. Organism_taxid: 1895

Biol. unit:

Tetramer (from PDB file)

Resolution:

1.40Å R-factor: 0.206 R-free: 0.221

Authors:

B.A.Katz

Key ref:

B.A.Katz (1997). Binding of biotin to streptavidin stabilizes intersubunit salt bridges between Asp61 and His87 at low pH. J Mol Biol, 274, 776-800. PubMed id: 9405158 DOI: 10.1006/jmbi.1997.1444

Date:

11-Sep-97 Release date: 14-Oct-98

Protein chain

P22629  (SAV_STRAV) -  Streptavidin from Streptomyces avidinii

Seq: 183 a.a.

Struc: 123 a.a.

DOI no: 10.1006/jmbi.1997.1444

J Mol Biol 274:776-800 (1997)

PubMed id: 9405158

Binding of biotin to streptavidin stabilizes intersubunit salt bridges between Asp61 and His87 at low pH.

B.A.Katz.

ABSTRACT

The remarkable stability of the streptavidin tetramer towards subunit dissociation becomes even greater upon binding of biotin. At two equivalent extensive monomer-monomer interfaces, monomers tightly associate into dimers that in turn associate into the tetramer at a less extensive dimer-dimer interface. To probe the structural basis for the enhancement of the stability of streptavidin by biotin, the crystal structures of apostreptavidin and its complexes with biotin and other small molecule and cyclic peptide ligands were determined and compared at resolutions as high as 1.36 A over a range of pH values from as low as 1.39. At low pH dramatic changes occur in the conformation and intersubunit hydrogen bonds involving the loop comprising Asp61 to Ser69. The hydrogen-bonded salt bridge between Asp61 Odelta2 and His87 Ndelta1, observed at higher pH, is replaced with a strong hydrogen bond between Asp61 Odelta1 and Asn85 Odelta1. Through crystallography at multiple pH values, the pH where this conformational change occurs, and thus the pKa of Asp61, was determined in crystals of space group I222 and/or I4122 of apostreptavidin and complexes. A range in pKa values for Asp61 was observed in these structures, the lowest being 1.78+/-0.19 for I222 streptavidin-biotin in 2.9 M (NH4)2SO4. At low pH the decrease in pKa of Asp61 and preservation of the intersubunit Asp61 Odelta2-Ndelta1 His87 hydrogen-bonded salt bridge in streptavidin-biotin versus apostreptavidin or streptavidin-peptide complexes is associated with an ordering of the flexible flap comprising residues Ala46 to Glu51, that in turn orders the Arg84 side-chain of a neighboring loop through resulting hydrogen bonds. Ordering of Arg84 in close proximity to the strong intersubunit interface appears to stabilize the conformation associated with the Asp61 Odelta2-Ndelta1 His87 hydrogen-bonded salt bridge. Thus, in addition to the established role of biotin in tetramer stabilization by direct mediation of intersubunit interactions at the weak interface through contact with Trp120, biotin may enhance tetramer stability at the strong interface more indirectly by ordering loop residues.

Selected figure(s)

Figure 1.
Figure 1. Structure of the I222 streptavidin-biotin tetramer, pH 2.00, showing the strong and weak intersubunit interfaces. The β-strands are colored yellow, the loop bearing Asp61 purple, the flexible flap that interacts with biotin green, the loop bearing Arg84 dark blue, and the remaining loops cyan. The Trp120 side-chain that interacts with biotin at the weak interface is shown. The His87 and Asp61 side-chains at the strong interface are shown in conformations associated with the intersubunit hydrogen-bonded salt bridge. Interloop hydrogen bonds involving Arg84, Glu51, and Asn49 are shown.

Figure 4.
Figure 4. (a) Superposition of the (2|F[o]|−|F[c]|), α[c] map onto the refined structure of I222 streptavidin-biotin, pH 2.00, 1.36 Å resolution, showing the binding site of biotin and its interaction with Trp120 of a neighboring subunit. Residues hydrogen bonding to biotin are labeled in yellow font. (b) Superposition of the (2|F[o]|−|F[c]|), α[c] map onto the refined structure of I222 streptavidin-2-iminobiotin, pH 3.25, 1.39 Å resolution. Residues hydrogen bonding to the ligand are labeled in yellow. Tyr43 is discretely disordered between two well-defined conformations involving a rotation of 5° in χ1. (c) Superposition of the (2|F[o]|−|F[c]|), α[c] map onto the refined structure of I222 streptavidin-glycoluril, pH 2.50, 1.40 Å resolution. Residues hydrogen bonding to the ligand are labeled in yellow. Normal hydrogen bonds mediating ligand binding are shown in yellow, and the NH → πTrp108 hydrogen bond in white. Note that Leu110 is discretely disordered.

The above figures are reprinted by permission from Elsevier: J Mol Biol (1997, 274, 776-800) copyright 1997.

Figures were selected by an automated process.