Refinement of triclinic lysozyme: I. Fourier and least-squares methods.

X-ray diffraction data to 1.5 A resolution have been collected for triclinic crystals of hen egg white lysozyme. The triclinic model was derived from the tetragonal one by the rotation function and refined initially by Fo-Fc and differential difference syntheses against 2 A resolution data. Refinement was continued by differential difference cycles against the 1.5 A data until R was reduced to 0.220. Although the initial refinement was rapid, it was subsequently a matter of attrition, leading to a complete recheck of the data and the discovery of systematic error which affected primarily the high-resolution data. Refinement was continued against the corrected 2 A data by block-diagonal least squares. After five cycles the refinement was terminated at R = 0.254 because of the imminent availability of a preferred refinement program. Problems with the protein model, the solvent, and the interaction of the scale and thermal parameters are discussed. The experiences gained in this study are summarized.

Unexpected similarities in the crystal structures of the Mcg light-chain dimer and its hybrid with the Weir protein.

The covalently linked hybrid of two human lambda-type light chains (Mcg and Weir) crystallizes as trigonal bipyramids in ammonium sulfate [Ely et al., Molec. Immun. 22, 85-92 (1985)]. While markedly different in appearance from the barrel-shaped crystals of the parental Mcg dimer, the bipyramids of the hybrid have the same space group: trigonal P3(1)21. Moreover, the unit cell dimensions are practically identical: a = 72.3 A in both proteins; c = 188.1 A in the hybrid and 185.9 A in the Mcg dimer. These observations imply that the crystal packing and the main features of the three-dimensional structures are closely similar in the Mcg X Weir hybrid and the Mcg dimer. The "constant" domains of the Mcg and Weir proteins belong to the same genetic subclass and were expected to interact in comparable ways in hybrids and parental dimers. However, the overall similarities in the "variable" domain pairs in the hybrid and Mcg dimer were completely unpredicted, since the amino acid sequences of the heterologous variable domains differ by 36 residues. By difference Fourier analysis the Weir light chain has been tentatively identified as monomer 1 (heavy-chain analogue) and the Mcg protein as monomer 2 (light-chain analogue) in the hybrid dimer. Substitutions in key positions in the hypervariable loops explain the differences in binding activity of the Mcg and Weir dimers. In the Mcg dimer bis(dinitrophenyl)lysine spans two relatively spacious subsites (A and B), with primary contacts involving tyrosines 34 and 38 of monomer 2. The Weir dimer, which does not bind dinitrophenyl ligands, has serine and phenylalanine in homologous positions. Moreover, the bilateral replacement of valine 48 and serine 91 in Mcg by leucine and methionine in the Weir dimer should effectively block access to subsite B. In the hybrid binding activity for bis(dinitrophenyl)lysine is restored because the Mcg light chain is present as the monomer 2 subunit.