REPARTITIONING OF NACL AND PROTEIN IMPURITIES IN LYSOZYME CRYSTALLIZATION

Nonuniform precipitant and impurity incorporation in protein crystals can cause lattice strain and, thus, possibly decrease the X-ray diffra ction resolution. To address this issue, a series of crystallization e xperiments were carried out, in which initial supersaturation, NaCl co ncentration, protein purity level and crystallized fraction were varie d. Lysozyme and protein impurities, as well as sodium and chloride wer e independently determined in the initial solution, supernatant and cr ystals. The segregation coefficients for Na+ and Cl- were found to be independent of supersaturation and NaCl concentration, and decreased w ith crystallized fraction/crystal size. Numerical evaluation of the ex tensive body of data, based on a nucleation-growth-repartitioning mode l, suggests a core of similar to 40 mu m in which salt is incorporated in much greater concentrations than during later growth. Small crysta ls containing higher amounts of incorporated NaCl also had higher prot ein impurity contents. This suggests that the excess salt is associate d with the protein impurities in the core. X-ray topography revealed s train fields in the center of the crystals comparable in size to the i nferred core. The growth rates of crystals smaller than 30-40 mu m in size were consistently 1.5-2 times lower than those of larger crystals , presumably due to higher chemical potentials in the core.