Heavy-atom sulphydryl derivatives of ox haemoglobin and β lactoglobulin; factors affecting isomorphism of native and derivative crystals

Abstract

The methods of preparing isomorphous heavy-atom derivatives of proteins are summarized and the advantages of reagents specific for particular amino acids are outlined. The coupling of mercurial reagents with the sulphydryl group of cysteine side chains is examined for the preparation of crystalline heavy-atom derivates of ox haemoglobin and β-lactoglobulin. Derivatives have been prepared with heavy atoms separated from the sulphydryl group by aromatic spacer groups. Matching ‘light-atom’ derivatives of ox haemoglobin have been prepared when distortion of the lattice occurred for the heavy-atom derivative. By preparing a variety of mercurial derivatives of ox haemoglobin using several reagent solvents it has been possible to trace factors which may influence isomorphism. Coupling of some of the same mercurials to β-lactoglobulin has been carried out and the necessary modifications in procedure are outlined together with differences in the character of derivatives produced. The general implications of the experiments for isomorphous replacement using specific amino acid reagents are discussed.