1a2y
From Proteopedia
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HEN EGG WHITE LYSOZYME, D18A MUTANT, IN COMPLEX WITH MOUSE MONOCLONAL ANTIBODY D1.3
Overview
Alanine scanning mutagenesis, double mutant cycles, and X-ray, crystallography were used to characterize the interface between the, anti-hen egg white lysozyme (HEL) antibody D1.3 and HEL. Twelve out of the, 13 nonglycine contact residues on HEL, as determined by the, high-resolution crystal structure of the D1.3-HEL complex, were, individually truncated to alanine. Only four positions showed a, DeltaDeltaG (DeltaGmutant - DeltaGwild-type) of greater than 1.0 kcal/mol, with HEL residue Gln121 proving the most critical for binding (DeltaDeltaG, = 2.9 kcal/mol). These residues form a contiguous patch at the periphery, of the epitope recognized by D1.3. To understand how potentially, disruptive mutations in the antigen are accommodated in the D1.3-HEL, interface, we determined the crystal structure to 1.5 A resolution of the, complex between D1.3 and HEL mutant Asp18 --> Ala. This mutation results, in a DeltaDeltaG of only 0.3 kcal/mol, despite the loss of a hydrogen bond, and seven van der Waals contacts to the Asp18 side chain. The crystal, structure reveals that three additional water molecules are stably, incorporated in the antigen-antibody interface at the site of the, mutation. These waters help fill the cavity created by the mutation and, form part of a rearranged solvent network linking the two proteins. To, further dissect the energetics of specific interactions in the D1.3-HEL, interface, double mutant cycles were carried out to measure the coupling, of 14 amino acid pairs, 10 of which are in direct contact in the crystal, structure. The highest coupling energies, 2.7 and 2.0 kcal/mol, were, measured between HEL residue Gln121 and D1.3 residues VLTrp92 and VLTyr32, respectively. The interaction between Gln121 and VLTrp92 consists of three, van der Waals contacts, while the interaction of Gln121 with VLTyr32 is, mediated by a hydrogen bond. Surprisingly, however, most cycles between, interface residues in direct contact in the crystal structure showed no, significant coupling. In particular, a number of hydrogen-bonded residue, pairs were found to make no net contribution to complex stabilization. We, attribute these results to accessibility of the mutation sites to water, such that the mutated residues exchange their interaction with each other, to interact with water. This implies that the strength of the, protein-protein hydrogen bonds in these particular cases is comparable to, that of the protein-water hydrogen bonds they replace. Thus, the simple, fact that two residues are in direct contact in a protein-protein, interface cannot be taken as evidence that there necessarily exists a, productive interaction between them. Rather, the majority of such contacts, may be energetically neutral, as in the D1.3-HEL complex.
About this Structure
1A2Y is a Protein complex structure of sequences from Gallus gallus and Mus musculus with PO4 as ligand. Active as Lysozyme, with EC number 3.2.1.17 Full crystallographic information is available from OCA.
Reference
A mutational analysis of binding interactions in an antigen-antibody protein-protein complex., Dall'Acqua W, Goldman ER, Lin W, Teng C, Tsuchiya D, Li H, Ysern X, Braden BC, Li Y, Smith-Gill SJ, Mariuzza RA, Biochemistry. 1998 Jun 2;37(22):7981-91. PMID:9609690
Page seeded by OCA on Sun Nov 18 09:24:58 2007
