Molecular dynamics of α-helical structure: poly-l-glutamic acid.

Communicated by Ramaswamy H. Sarma.

[Zn-CysHis Protein Factor Families: Role of Electrostatic Interaction of Zn-Domains in Factor Functions].

The large amount of the DNA- and RNA-binding protein regulatory factors contain Zn-domains. Total six Zn-CysHis factor families with different functions were considered. Among them the transcription factors of family PF00096, complex type Zn-Cys2His2, is the most important because it includes over 340.000 members of 365.000 total members listed at present in the Zn-containing factor families. The role of electrostatic potential of Zn-domains in the structure and function of factors from different factor families has been considered. We have shown that the positive electrostatic potential of Zn-domains plays an essential role in forming contact of factors' Zn-domains with DNA/RNA in three protein factor families PF00096, PF12874, and PF09329. While in other three families PF03119, PF08996, and PF01258 Zn-domains do not contact with nucleic acids, and electrostatic interactions do not play a distinctive role, consequently.

Side projections of double-helical DNA: example of binding patterns of DNA in the complex with factor TFIIIA.

Communicated by Ramaswamy H. Sarma.

The electrostatic role of the Zn-Cys2His2 complex in binding of operator DNA with transcription factors: mouse EGR-1 from the Cys2His2 family.

The mouse factor Zif268, known also as early growth response protein EGR-1, is a classical representative for the Cys2His2 transcription factor family. It is required for binding the RNA polymerase with operator dsDNA to initialize the transcription process. We have shown that only in this family of total six Zn-finger protein families the Zn complex plays a significant role in the protein-DNA binding. Electrostatic feature of this complex in the binding of factor Zif268 from Mus musculus with operator DNA has been considered. The factor consists of three similar Zn-finger units which bind with triplets of coding DNA. Essential contacts of the factor with the DNA phosphates are formed by three conservative His residues, one in each finger. We describe here the results of calculations of the electrostatic potentials for the Zn-Cys2His2 complex, Zn-finger unit 1, and the whole transcription factor. The potential of Zif268 has a positive area on the factor surface, and it corresponds exactly to the binding sites of each of Zn-finger units. The main part of these areas is determined by conservative His residues, which form contacts with the DNA phosphate groups. Our result shows that the electrostatic positive potential of this histidine residue is enhanced due to the Zn complex. The other contacts of the Zn-finger with DNA are related to nucleotide bases, and they are responsible for the sequence-specific binding with DNA. This result may be extended to all other members of the Cys2His2 transcription factor family.

Spatial sign-alternating charge clusters in globular proteins.

Large sign-alternating charge clusters formed by the charged side groups of amino acid residues and N- and C-terminal groups were found in the majority of considered globular proteins, namely 235 in a total of 274 protein structures, i.e. 85.8%. The clusters were determined by the criteria proposed earlier: charged groups were included in the cluster if their charged N and O atoms were located at distances between 2.4 and 7.0 A. The set of selected proteins consisted of known non-homologous protein structures from the Protein Data Bank with a resolution less than or equal to 2.5 A and pair sequence similarity less than 25%. Molecular masses of the proteins were from 5.5 to 91.5 kDa and protein chain length from 50 to 830 residues. The distribution of charged groups on the protein surface between isolated charged groups, small clusters with two and three groups, and large clusters with four or more groups were found to be approximately similar making 33, 35 and 32% of the total amount of protein charged groups, respectively. The large sign-alternating charge clusters with four or more charged groups were studied in greater detail. The amount of such clusters depends on the protein chain length. The small proteins contain 1-3 clusters while the large proteins display 4-6 or more clusters. On average, 1.5 clusters per each 100 residues were observed. In contrast with this, the size of a cluster, i.e. the number of charged groups inside a cluster, does not depend on the protein molecular mass, and large clusters are observed for proteins from a range of molecular masses. Clusters consisting of four to six charged groups occur most frequently, although extra large clusters are also often revealed. We can conclude that sign-alternating charge clusters are a common feature of the protein surface of globular protein. They are suggested to play a general functional role as a local polar factor of protein surface.

Alternating charge clusters of side chains: new surface structural invariants observed in calf eye lens gamma-crystallins.

A detailed stereochemical analysis of the oppositely charged side chains of amino acid residues on the surface of calf eye lens protein gamma-crystallin B has been carried out. The refined structural data of very high quality obtained at 1.47 A resolution have been used. Charge-charge interactions were considered to be valuable for all the charged oxygen and nitrogen atoms situated at distances, d, between 2.4 and 7.0 A. This means we consider short contact ion pairs as those with intercharge distances 2.4 < d < or = 4.0 A and distant contact ion pairs as those with distances 4.0 < d < or = 7.0 A. Hydrogen bonding of the charged atomic groups with the structural water molecules also has been considered. We have not looked at the side groups of histidines which are charged only partially at neutral pH. Five clusters of charged side chains which were large enough were observed. The clusters are comprised of four to six residues which compose 54.3% of the total charged residues in the protein. The clusters contain from eight to 12 charged atoms and look like the bent chains of oppositely charged atoms. All clusters are of plane geometry and their maximal linear dimensions are from 11 to 18 A. The root mean square deviations of charged atoms from the cluster plane varied from 0.63 to 0.86 A for four clusters and was only 1.85 A for the largest cluster. All clusters include a number of water molecules situated on the cluster boundary and grouped near the cluster plane. It was shown that the amino acid sequence positions of charged residues are conservative for all the proteins of the gamma-crystallin family of vertebrates including fish, frog, mouse, rat, calf and human. The cluster properties were discussed both in their functional aspect for gamma-crystallins and in other aspects common for globular proteins. As a result, the alternating charge clusters should be considered as newly recognized surface structural invariants. The importance of the charged side chain clusters is claimed for the updated concept of the protein surface.

Structure of bovine eye lens gammaD (gammaIIIb)-crystallin at 1.95 A.

The crystal structure of bovine lens gammaIIIb-crystallin at 2.5 A resolution previously reported was interpreted using a consensus sequence derived from related vertebrate sequences on the assumption that gammaIIIb-crystallin derived from the gammaC-crystallin gene. It has recently been shown that gammaIIIb is a product of the bovine gammaD gene. The structure of gammaIIIb has now been refined with the bovine gammaD sequence using new 1.95 A resolution synchrotron data. The crystallographic R factor was 20.4% for all 33 104 reflection data between 8.0 and 1.95 A measured at 277(1) K. The electron density fully supported the assignment of the gammaD sequence to gammaIIIb. The crystal belongs to space group P2(1)2(1)2(1) with two molecules of molecular mass 20 749 Da in the asymmetric unit in which 219 water molecules were located. The two-domain four-Greek-key motif highly symmetrical protein is very similar in structure to gammaB-crystallin (81% sequence identity). There is a single amino-acid deletion in gammaD in the linker region connecting the two domains. The intermolecular oganization in the crystal lattice is quite different from gammaB as a result of key mutations involving surface residues Leu51, Ile103 and His155. These point mutations will contribute to the intermolecular behaviour of the gamma-crystallins in the eye lens, where they are major components of the densely packed, high refractive index regions of the lens.

Surface interactions of gamma-crystallins in the crystal medium in relation to their association in the eye lens.

A comparative study of intermolecular interactions in crystals of two homologous low molecular weight proteins, gamma-II and gamma-IIIb crystallins, from calf eye lens was carried out. Crystal packings for these proteins are very different: intermolecular contact areas compose about 33% of the total accessible surface area of gamma-II as compared with 13% in gamma-III. Two key residues seem to be mainly responsible for the differences in protein association in the crystal medium. These are Ser 103 and Leu 155 in gamma-II, which are replaced by Met 103 and His 155 in gamma-IIb. A similar substitution of these residues is observed in different gene products of gamma-crystallins from a number of vertebrates. This is consistent with the existence of a genetically controlled mechanism for determining intermolecular association of gamma-crystallins in the native medium of the lens.

Quantitative study of secondary structure of histones H1, H2A, and H4 in solution by infrared spectroscopy.

The secondary structure of histones H1, H2A, and H4 (F1, F2a2, and F2a1) has been quantitatively studied in heavy water (2H2O) solutions in a wide range of histone concentration, p2H, and concentration of sodium chloride using an improved infrared spectroscopy method. Under all conditions there are about 5--10% of alpha helix. Conditions favourable for aggregation induce formation of antiparallel pleated sheet structure to an extent of about 15% in H1 and H2A and about 30% in H4. When the p2H and concentration of NaCl are in the physiological range, there is the same content of this structure in H2A and H4 and none in H1.