Biomedical subjects

H P Driessen

Publications and source records attributed to H P Driessen.

12 recordsLinked to original sources

PubMed1992

A model of the structure of human annexin VI bound to lipid monolayers.

Annexin VI is an eight repeat member of the annexin family of proteins which are both water soluble and bind to negatively charged phospholipids in a calcium-dependent manner. Here we present a model for annexin VI based on fitting the three-dimensional structure of two annexin V molecules (Huber (1990) EMBO J. 9, 3867-3874) to the two-dimensional stain-excluding density of lipid-bound annexin VI (Newman (1989) J. Mol. Biol. 206, 213-219). Both annexin VI lobes could only be fitted with their convex faces closest to the lipid monolayer. This supports the hypothesis that annexin-lipid binding is mediated by the interaction between calcium bound to the loops protruding from the convex protein surface and phospholipid headgroups.

Annexin A6↗

PubMed1992

X-ray analyses of peptide-inhibitor complexes define the structural basis of specificity for human and mouse renins.

X-ray analyses have defined the three-dimensional structures of crystals of mouse and human renins complexed with peptide inhibitors at resolutions of 1.9 and 2.8 A, respectively. The exquisite specificity of renin arises partly from ordered loop regions at the periphery of the binding cleft. Although the pattern of main-chain hydrogen bonding in other aspartic proteinase inhibitor complexes is conserved in renins, differences in the positions of secondary structure elements (particularly helices) also lead to improved specificity in renins for angiotensinogen substrates.

Amino Acid Sequence↗

PubMed1991

Structure of oligomeric beta B2-crystallin: an application of the T2 translation function to an asymmetric unit containing two dimers.

The molecular structure of the main subunit of the beta-crystallins, components of the vertebrate eye lens, has recently been solved by molecular replacement at 2.1 A resolution [Bax, Lapatto, Nalini, Driessen, Lindley, Mahadevan, Blundell & Slingsby (1990). Nature (London), 347, 776-780]. The protein, beta B2, is a dimer in solution, but a tetramer in the crystal with one subunit in the asymmetric unit of space group I222. Using the crystallographic dimer from this I-centred form the structure of a C222 crystal form of the beta B2 protein with four subunits in the asymmetric unit has now been solved by molecular replacement at 3.3 A. The solution involved the use of a new translation function for non-crystallographic symmetry, based on the T2 function of Crowther & Blow [Acta Cryst. (1967), 23, 544-548].

Crystallins↗

PubMed1991

Isolation and characterization of cDNAs encoding beta A2- and beta A4-crystallins: heterologous interactions in the predicted beta A4-beta B2 heterodimer.

Except for the two acidic chains, beta A2 and beta A4, the primary structures of all bovine beta-crystallins have previously been elucidated, either by direct protein sequencing or prediction from cDNA sequencing. Both beta A2 and beta A4 were found to be synthesized in half-year-old calf lenses and are therefore likely to be present in a cDNA bovine library constructed from mRNA isolated from lenses of that age. A large number of cDNA clones was screened with all available crystallin, actin, vimentin and lens membrane protein MP26 probes and finally with a randomly primed mRNA probe. Clones positive for the latter, but negative for known lens proteins, were isolated and sequenced. beta A2, comprising 197 aa, and beta A4, comprising 209 aa, were identified. Both proteins have a conserved two-domain structure and an N-terminal extension which is variable. A three-dimensional model of the structure of beta A4 was made based on the coordinates of one subunit from the beta B2 dimer which has recently been solved using x-ray diffraction techniques. The resulting heterodimer structure, together with the compiled bovine beta-crystallin sequences, was used to indicate those regions of the sequences which distinguish acidic from basic beta-crystallins with a view to defining structural features necessary for subunit recognition in beta-crystallin aggregates. With the aid of the present data, the complete evolutionary tree of the bovine beta-crystallin family has been constructed, which confirms the early separation of the genes encoding the three acidic and the three basic beta-crystallins.

Amino Acid Sequence↗

PubMed1991

MOLPACK: molecular graphics for studying the packing of protein molecules in the crystallographic unit cell.

A graphics program, MOLPACK, has been developed on the Silicon Graphics IRIS-4D computer system for displaying the packing of proteins in the crystallographic unit cell. In addition to the normal viewing operations of rotation, translation and scaling, the program has the ability to translate molecules along the cell axes while maintaining their crystallographic equivalent positions within the unit cell. This allows the user to observe the packing of protein molecules generated by molecular replacement, to create a new packing model or to locate an unknown molecule. A special feature of the program is that up to four independent molecules can be manipulated in the asymmetric unit.

Computer Graphics↗

PubMed1990

Crystallization and preliminary X-ray crystallographic studies of human placental annexin IV.

Human placental annexin IV, a member of the annexin family of calcium and phospholipid-binding proteins, has been crystallized by the vapour diffusion method in the presence of calcium, using polyethylene glycol 8000. The crystals are orthorhombic, space C222(1), cell dimensions a = 105.4 A, b = 115.7 A, c = 80.7 A and diffract to at least 2.5 A resolution on a synchrotron source.

Binding Sites↗

PubMed1990

The E2 protein of human papillomavirus type 16. Over-expression and purification of an active transcriptional regulator.

The E2 open reading frame of human papillomavirus type 16 was inserted into the Escherichia coli vector pKK223-3, and expressed to greater than 15% of total cellular protein when induced with isopropyl beta-D-thiogalactopyranoside. The highest expressing clone was grown in bulk and the E2 protein purified to homogeneity by the following procedure: (a) isolation of the insoluble protein fraction; (b) extraction with urea; (c) quaternary amino-ethyl-Sepharose ion-exchange chromatography and (d) renaturation and chromatography on dextran sulphate. That the purified protein was fully functionally active was confirmed by its specific DNA-binding properties and its ability to activate gene transcription by over two orders of magnitude in an in vivo assay.

Base Sequence↗

PubMed1989

Structure of the bovine eye lens gamma s-crystallin gene (formerly beta s).

The organization of a number of crystallin genes has already been resolved. One of the remaining genes of which the structure was hitherto unknown is the gamma s gene (formerly beta s). We determined the complete sequence of the bovine gamma s-crystallin-coding gene, apart from the middle region of the first intron. Since it contains three exons and two introns, we conclude that the former beta s, also at the gene level is gamma-crystallin-like. However, it is located on chromosome 3, in contrast to other gamma genes which occur in tandem on the human chromosome 2.

Amino Acid Sequence↗

PubMed1989

Packing interactions in the eye-lens. Structural analysis, internal symmetry and lattice interactions of bovine gamma IVa-crystallin.

gamma-Crystallins are a family of low molecular weight proteins found in high concentration in the densely packed regions of high refractive index in vertebrate lenses. Certain members have the characteristic property of a high critical temperature (tc) for phase separation. We report the three-dimensional structure determination of such a protein, bovine lens gamma IVa-crystallin, which has been refined to give an X-ray R-factor of 0.143. Its high tc contrasts with the low tc gamma II-crystallin, whose structure we have already published. The root mean square difference between the alpha-carbon atoms of these two proteins is 0.70 A and gamma IVa has an internal symmetry even higher than that of gamma II. The presence of a protein that exhibits the phenomenon of phase separation at body temperature renders the lens very susceptible to a transformation from transparent to an opaque state due to irregularities in the refractive index. Protein interactions of gamma IVa-crystallin have implications for the mechanism of cataract formation. Modes of self-association behaviour of gamma IVa-crystallin have been inferred from an analysis of the lattice interactions in the crystalline state, where the protein packing density is similar to that of the intact lens. It appears that the point mutation at position 103 from a serine residue in gamma II to a valine in gamma IVa gives rise to a lattice contact formed by two four-stranded beta-sheets in gamma IVa. A group-specific mutation at position 118 from leucine to phenylalanine induces subtle differences in core packing, leading to a reorganization around residue 103. However, the final phase separation determinant may be a complex combination of many side-chain functions.

Amino Acid Sequence↗

PubMed1988

The use of pseudosymmetry in the rotation function of gamma IVa-crystallin.

Bovine lens gamma IVa-crystallin crystallizes in space group C222(1) with cell dimensions a = 35.1, b = 46.2, c = 186.2 A, and contains one molecule in the asymmetric unit. The structure was determined at 3.0 A resolution using cross-rotation functions and R-factor searches with the bovine lens protein gamma II-crystallin as the model structure. The rotation function appears to be very sensitive to the resolution range and type of coefficient employed; the use of normalized structure-factor amplitudes gave the best results. The potential problem of a pseudo solution due to an internal pseudo-twofold axis was put to advantage by aligning this axis parallel to z. The results of the R-factor search were well defined. The molecular replacement solution was improved by rigid-body least-squares refinement, initially of the whole molecule, then for the two domains. The R factor at this stage was 39.4% at 2.3-10.0 A. The gamma IVa structure has an even higher internal symmetry than gamma II, since the two domains are related by a rotation around the pseudo-twofold axis of 178.7 degrees as compared with 176.2 degrees for gamma II.

Animals↗

PubMed1988

Evolutionary and functional relationships between the basic and acidic beta-crystallins.

beta-Crystallins are complex oligomers composed of many related subunits. In order to understand their interactions we have built molecular models of several bovine beta-crystallins, based on their sequence similarity to the well-defined gamma-II crystallin structure, using interactive computer graphics techniques. Their common origin with gamma-crystallin is displayed in both the retention of four-fold sequence repeats of critical residues involved with stabilizing a folded beta-hairpin and the conservation of core-filling hydrophobic side-chains. The beta-crystallins have been built as bilobal molecules with each domain composed of two 'Greek key' motifs which associate about an approximate two-fold axis to form beta-sheets. The beta-crystallin sequences have previously been shown to comprise two families, the basic and acidic subunits, which have extensions of sequence. The three-dimensional models show how the two families appear to stabilize the folded beta-hairpin in the N- and C-terminal domains in ways which suggest that they have diverged from a common ancestor in different ways. Acidic beta-crystallins, like gamma-crystallins, have a regular array of charges on their N-terminal domain which has been interrupted in basic beta-crystallins by hydrophobic residues which may be related to the presence of a C-terminal extension. beta-Crystallins are more highly charged than gamma-crystallins although their charge density is higher in certain regions of the N-terminal domain, particularly in beta B1-crystallin. beta-crystallins also differ from gamma-crystallins in the virtual absence of core-filling sulphydryl groups whereas they have numerous sulphur-containing side-chains together with tryptophan and histidine rings protruding from the globular domains, particularly in the acidic subunits. The burial of these residues in subunit contacts is consistent with their spectroscopic and electrostatic properties. Protein subunit aggregation commonly occurs through hydrophobic interaction or beta-sheet extension. Analysis of the subunit surfaces has identified an N-terminal hydrophobic region common to beta B1 and beta B2 whereas a C-terminal hydrophobic loop region is common to beta B1 and beta A1 and may be correlated with their association properties. It is suggested that the polar C-terminal domain of beta B2 contributes towards the solubility of higher aggregates by interactions involving beta-sheet structure.

Amino Acid Sequence↗

PubMed1985

The mechanism of N-terminal acetylation of proteins.

N alpha-acetylation is almost exclusively restricted to eukaryotic structural proteins. As a rule it is a post-initiational process, requiring the presence of the enzyme N alpha-acetyltransferase and the acetyl donor acetylcoenzyme A. N alpha-acetyltransferases appear to have a narrow substrate specificity, which is very similar for enzymes from different tissues and species. Amino acids predominantly present at the N terminus of N alpha-acetylated proteins are alanine, serine, and methionine. The occurrence of these residues is apparently a prerequisite for acetylation. The region following these amino acids is also important. If methionine is at the N terminus, the second position is always occupied by a strongly hydrophilic amino acid. Two- and three-dimensional structural characteristics of the protein do not seem to play a major role in N alpha-acetylation. Up to now the exact function for N alpha-acetylation is not known.

Acetylation↗