The crystallography of calf rennin (chymosin)

Abstract

The growth of crystals of calf rennin (chymosin, † EC 3.4.4.3) and the control of nucleation to produce crystals of desired size, are described. Only one stable species, orthorhombic rennin I, has been found, but a metastable monoclinic species, rennin II, appeared on one occasion in a solution nearly saturated with glycine. The morphology, optical characteristics and unit cells of these species are recorded. In rennin I, small differences in the properties of sectors built by deposition on the three types of crystal faces are attributed to differences in the proportions of either degradation products or of the slightly different isoenzymes known to be present in calf rennin. Rennin I crystals have been obtained only within the pH range of greatest stability of the protein, 5. 0 to 6. 2; but the crystals tolerate subsequent changes of pH down to 2. 0. The stability of crystals appears to be much greater than that of salt-free protein solutions. Rennin I may be crosslinked with glutaraldehyde with no loss of order, giving crystals stable over a wide range of solution conditions, including pH 2.0 to 10.0 and salt-free solutions. The remarkable swelling behaviour when the pH is raised beyond the stable range is described. Rennin II, though monoclinic in symmetry, shows in its X -ray diffraction patterns strong evidence of a pseudo-orthorhombic arrangement of molecules.