In situ atomic force microscopy studies of surface morphology, growth kinetics, defect structure and dissolution in macromolecular crystallization

Surface morphologies of thaumatin, catalase, lysozyme and xylanase crystals were investigated using in situ atomic force microscopy. For thaumatin, lysozyme and xylanase crystals, growth steps having a height equal to the unit cell parameter were produced both by screw dislocations and two-dimensional nuclei. Growth of catalase crystals proceeded in alternating patterns exclusively by two-dimensional nucleation and the successive deposition of distinctive growth layers, each having a step height equal to half the unit cell parameter. The shapes of islands on successive layers were related by 2-fold rotation axes along the [0 0 1] direction. Experiments revealed that step bunching on crystalline surfaces occurred either due to two- or three-dimensional nucleation on the terraces of vicinal slopes or as a result of uneven step generation by complex dislocation sources. Growth kinetics for thaumatin and catalase crystals were investigated over wide supersaturation ranges. Strong directional kinetic anisotropy in the tangential step growth rates in different directions was seen. From the supersaturation dependencies of tangential step rates and the rates of two-dimensional nucleation, the kinetic coefficients of the steps and the surface free energy of the step edge were calculated. Adsorption of impurities which formed filaments on the surfaces of catalase and thaumatin crystals was recorded. Cessation of growth of xylanase and lysozyme crystals was also observed and appeared to be a consequence of the formation of dense impurity adsorption layers. Crystal growth resumed upon scarring of the impurity adsorption layer and clearing of the crystal surface with the AFM tip. Adsorption of three-dimensional clusters, which consequently developed into either properly aligned multilayer stacks or misaligned microcrystals was recorded. For catalase crystals, incorporation of misoriented microcrystals as large as 50 x 3 x 0.1 mu m(3) produced elastic deformations in growth layers of approximate to 0.6%, but did not result in the defect formation. Etching experiments on catalase crystals revealed high defect densities. (C) 1999 Elsevier Science B.V. All rights reserved.