[Study of crystals of aspartate aminotransferase complexed with D-aspartate]. / Issledovanie kristallov kompleksa aspartat-aminotransferazy s D-aspartatom.

We report here the first X-ray studies of the complex of cytosolic aspartate aminotransferase from chicken heart with D-aspartate at 2.5 A resolution. Crystals of the complex were grown by cocrystallization (space group is P2(1)2(1)2(1), parameters: a = 62.48 A, b = 117.71 A, c = 124.38 A). They contain one dimeric molecule in the asymmetric unit. The X-ray analysis proves that attachment of D-aspartate induces considerable conformational changes in the active sites of two subunits of the enzyme: both subunits of the complex are in the closed conformation, the interaction of the enzyme with D-aspartate induces a substantial turn (about 90 degrees) of the coenzyme in one subunit, the coenzyme ring is deformed, considerable conformational changes are determined for Phe-18 and Glu-141. Apparently, the amino group of the substrate is a trigger of the conformational changes in the active site of the enzyme.

Modulation of calmodulin plasticity in molecular recognition on the basis of x-ray structures.

Calmodulin is the primary calcium-dependent signal transducer and regulator of a wide variety of essential cellular functions. The structure of calcium-calmodulin bound to the peptide corresponding to the calmodulin-binding domain of brain calmodulin-dependent protein kinase II alpha was determined to 2 angstrom resolution. A comparison to two other calcium-calmodulin structures reveals how the central helix unwinds in order to position the two domains optimally in the recognition of different target enzymes and clarifies the role of calcium in maintaining recognition-competent domain structures.

Calmodulin structure refined at 1.7 A resolution.

We have determined and refined the crystal structure of a recombinant calmodulin at 1.7 A resolution. The structure was determined by molecular replacement, using the 2.2 A published native bovine brain structure as the starting model. The final crystallographic R-factor, using 14,469 reflections in the 10.0 to 1.7 A range with structure factors exceeding 0.5 sigma, is 0.216. Bond lengths and bond angle distances have root-mean-square deviations from ideal values of 0.009 A and 0.032 A, respectively. The final model consists of 1279 non-hydrogen atoms, including four calcium ions, 1130 protein atoms, including three Asp118 side-chain atoms in double conformation, 139 water molecules and one ethanol molecule. The electron densities for residues 1 to 4 and 148 of calmodulin are poorly defined, and not included in our model, except for main-chain atoms of residue 4. The calmodulin structure from our crystals is very similar to the earlier 2.2 A structure described by Babu and coworkers with a root-mean-square deviation of 0.36 A. Calmodulin remains a dumb-bell-shaped molecule, with similar lobes and connected by a central alpha-helix. Each lobe contains three alpha-helices and two Ca2+ binding EF hand loops, with a short antiparallel beta-sheet between adjacent EF hand loops and one non-EF hand loop. There are some differences in the structure of the central helix. The crystal packing is extensively studied, and facile crystal growth along the z-axis of the triclinic crystals is explained. Herein, we describe hydrogen bonding in the various secondary structure elements and hydration of calmodulin.

Target enzyme recognition by calmodulin: 2.4 A structure of a calmodulin-peptide complex.

The crystal structure of calcium-bound calmodulin (Ca(2+)-CaM) bound to a peptide analog of the CaM-binding region of chicken smooth muscle myosin light chain kinase has been determined and refined to a resolution of 2.4 angstroms (A). The structure is compact and has the shape of an ellipsoid (axial ratio approximately 2:1). The bound CaM forms a tunnel diagonal to its long axis that engulfs the helical peptide, with the hydrophobic regions of CaM melded into a single area that closely covers the hydrophobic side of the peptide. There is a remarkably high pseudo-twofold symmetry between the closely associated domains. The central helix of the native CaM is unwound and expanded into a bend between residues 73 and 77. About 185 contacts (less than 4 A) are formed between CaM and the peptide, with van der Waals contacts comprising approximately 80% of this total.

Novel subunit structure observed for noncooperative hemoglobin from Urechis caupo.

Tetrameric hemoglobin from the "fat innkeeper" worm Urechis caupo possesses a novel subunit arrangement having an "inside out" quaternary structure in that the G/H helices are located on the outer surface of the tetramer. A 5-A resolution crystal structure reveals that although the individual subunits are beta-like, having a distinct D helix and the general myoglobin fold, the subunit contacts are very different from those previously observed for hemoglobins. Furthermore, the hemoglobin from U. caupo is also quite different from the unusual hemoglobin tetramer from clam which also has its G/H helices on the outer surface but with the hemes in close proximity through E-F helical contacts (Royer, W. E., Jr., Love, W. E., and Fenderson, F. F. (1985) Nature 316, 277-280).

Crystallization and subunit structure of histidine decarboxylase from Lactobacillus 30a.

Histidine decarboxylase from Lactobacillus 30a has been crystallized in a variety of forms which together indicate a revised subunit structure for the native particle. Octahedral crystals of the wild type enzyme obtained at room temperature from ammonium sulfate solutions in microdiffusion cells belong to tetragonal space group I4122 with a = b = 222 A and c = 107.5 A. Trigonal and hexagonal plates of prohistidine decarboxylase and activated proenzyme obtained at 4 degrees C from polyethyleneglycol solutions by vapor equilibration using the hanging drop technique belong to the trigonal space group P321 with a = b = 100 A and c = 164 A. The space group symmetries and unit cell contents of these crystals indicate 32 point group symmetry for the subunit structure of these enzymes. Sedimentation coefficients of wild type enzyme measured as a function of ionic strength at pH 7.0 indicate a rapid equilibrium between species varying from 6.9 S to 9.4 S. Sedimentation equilibrium analysis demonstrated the existence of a nearly homogeneous particle with Mr congruent to 208,000 at ionic strengths above I = 0.20, while an additional species of approximately one-half that molecular weight is observed at very ionic strengths (I = 0.2). At the pH optimum of the enzyme (pH 4.8), te larger species is dominant at all ionic strengths tested. Electron micrographs of native wild type enzyme show a dominant tetrahedral particle approximately 60 A on an edge while similar micrographs of enzyme cross-linked with glutaraldehyde show a dumbbell-shaped particle approximately 60 A in width and 120 A in length. These results establish that: (a) the native enzyme has a Mr congruent to 208,000 and a subunit composition (alpha beta)6; (b) the proenzyme has a subunit composition (pi)6; and (c) stable (alpha beta)3 and (pi) 3 particles exist under certain conditions.

Diffusion approximation for large absorption in radiative transfer.

A new diffusion model is developed for radiative transfer in particulate media. It includes the effects of higher Legendre moments while avoiding the mathematical complexities of solving multiple coupled differential moment equations and satisfying higher-order boundary conditions. The method accurately extrapolates the conventional Eddington approximation to problems involving large absorption. Although the simplifying assumptions limit the model to nonbeam incidence and spatial homogeneity of the scattering material, they are nonrestrictive with regard to the size and shape of the medium, the character of background reflections, and the type of phase function.

Six-beam models in radiative transfer theory.

A critical analysis is given of the applicability of six-beam models to radiative transfer in particulate materials. The method of introducing transverse scattering in these models is shown to cause fundamental difficulties in the case of physically plausible phase functions; in particular, the effective absorptivity is abnormally large and thus results in incorrect reflectances and transmittances. Six-beam calculations for several media are compared with accurate solutions, with Chu-Churchill two-beam results, and with a simple modification to the Eddington approximation, the last being generally superior over a wide range of conditions.