Biomedical subjects

M Hennig

Publications and source records attributed to M Hennig.

51 records · Page 3Linked to original sources

PubMed1995

2.0 A structure of indole-3-glycerol phosphate synthase from the hyperthermophile Sulfolobus solfataricus: possible determinants of protein stability.

BACKGROUND: Recent efforts to understand the basis of protein stability have focused attention on comparative studies of proteins from hyperthermophilic and mesophilic organisms. Most work to date has been on either oligomeric enzymes or monomers comprising more than one domain. Such studies are hampered by the need to distinguish between stabilizing interactions acting between subunits or domains from those acting within domains. In order to simplify the search for determinants of protein stability we have chosen to study the monomeric enzyme indole-3-glycerol phosphate synthase from the hyperthermophilic archaeon Sulfolobus solfataricus (sIGPS), which grows optimally at 90 degrees C. RESULTS: The 2.0 A crystal structure of sIGPS was determined and compared with the known 2.0 A structure of the IGPS domain of the bifunctional enzyme from the mesophilic bacterium Escherichia coli (eIGPS). sIGPS and eIGPS have only 30% sequence identity, but share high structural similarity. Both are single-domain (beta/alpha)8 barrel proteins, with one (eIGPS) or two (sIGPS) additional helices inserted before the first beta strand. The thermostable sIGPS has many more salt bridges than eIGPS. Several salt bridges crosslink adjacent alpha helices or participate in triple or quadruple salt-bridge clusters. The number of helix capping, dipole stabilizing and hydrophobic interactions is also increased in sIGPS. CONCLUSIONS: The higher stability of sIGPS compared with eIGPS seems to be the result of several improved interactions. These include a larger number of salt bridges, stabilization of alpha helices and strengthening of both polypeptide chain termini and solvent-exposed loops.

Aldehyde Oxidoreductases↗

PubMed1995

Crystal structure of concanavalin B at 1.65 A resolution. An "inactivated" chitinase from seeds of Canavalia ensiformis.

Seeds of Canavalia ensiformis (jack bean) contain besides large amounts of canavalin and concanavalin A, a protein with a molecular mass of 33,800 which has been named concanavalin B. Although concanavalin B shares about 40% sequence identity with plant chitinases belonging to glycosyl hydrolase family 18, no chitinase activity could be detected for this protein. To resolve this incongruity concanavalin B was crystallised and its three-dimensional structure determined at 1.65 A (1 A = 0.1 nm) resolution. The structure consists of a single domain with a (beta/alpha)8 topology. A 30 amino acid residue long loop occurs between the second beta-strand of the barrel and the second alpha-helix. This extended loop is unusual for the (beta/alpha)8 topology, but appears in a similar conformation in the structures of the seed protein narbonin and several chitinases as well. Two non-proline cis-peptide bonds are present in the structure of concanavalin B: Ser34-Phe, and Trp265-Asn. This structural feature is rarely observed in proteins, but could also be identified in the three-dimensional structures of family 18 chitinases and narbonin in coincident positions. In the chitinases the aromatic residues of the non-proline cis-peptides have been proposed to have a function in the binding of the substrate. The region in concanavalin B, where in chitinases the active site is located, shows two significant differences. First, the catalytic glutamic acid is a glutamine in concanavalin B. Second, although part of the substrate binding cleft of the chitinases is present in concanavalin B, it is much shorter. From this we conclude that concanavalin B and family 18 chitinases are closely related, but that concanavalin B has lost its enzymatic function. It still may act as a carbohydrate binding protein, however.

Amino Acid Sequence↗

PubMed1995

Crystal structure of narbonin at 1.8 A resolution.

The three-dimensional structure of narbonin, a seed protein from Vicia narbonensis L, has been determined at 1.8 A resolution. Phase information was obtained by multiple isomorphous replacement and optimized anomalous dispersion. The narbonin structure was initially traced with only 17% amino-acid sequence information and preliminarily refined to a crystallographic R-factor of 16.5%. It is now refined to 15.9% using full sequence information derived from cDNA and after the addition of more solvent molecules. The monomeric molecule of narbonin is an eight-stranded parallel beta-barrel surrounded by alpha-helices in a beta/alpha-topology similar to that first observed in triose phosphate isomerase. Differences exist in the N-terminal part of the polypeptide chain, where the first helix is replaced by a loop and the second beta-strand is followed by an additional antiparallel alpha-sheet placed parallel on top of alpha-helices alpha3 and alpha4. Two short additional secondary structures are present. The first, an alpha-helix, is situated between the seventh beta-strand and the following helix, and the second, which is a 3(10) helix, between the eighth strand and the C-terminal helix. The most striking observation is the lack of a known enzymatic function for narbonin, because all TIM-like structures known so far are enzymes.

Journal Article↗

PubMed1994

Crystallization and preliminary X-ray diffraction studies of recombinant human ornithine aminotransferase.

Human liver ornithine aminotransferase was expressed in Escherichia coli and purified by ammonium sulfate fractionation and anion exchange column chromatography. The purified recombinant enzyme is fully active and crystallized readily over a wide range of polyethylene glycol concentrations. The crystals belong to the trigonal space group P3(1)21 (or its enantiomorph P3(2)21) with unit cell parameters a = b = 116.3 A, and c = 190.0 A, alpha = beta = 90 degrees, gamma = 120 degrees. There are three monomers per asymmetric unit. Self-rotation function studies revealed both 2-fold and 3-fold non-crystallographic symmetry, with the local 3-fold axis being tilted 15 degrees from the c axis and perpendicular to a crystallographic dyad. A complete native data set to 2.3 A resolution was collected using synchrotron radiation.

Cloning, Molecular↗

PubMed1994

Crystallization and preliminary X-ray analysis of wild-type and K272A mutant glutamate 1-semialdehyde aminotransferase from Synechococcus.

Crystals of the pyridoxal-5'-phosphate dependent enzyme glutamate-1-semialdehyde aminotransferase (EC 5.4.3.8) from Synechococcus have been grown from polyethylene glycol solutions. The wild-type enzyme crystallizes in space group P2(1)2(1)2(1), with cell dimensions a = 68.4 A, b = 108.0 A, c = 122.6 A. The inactive mutant in which the cofactor-binding lysine 272 residue is replaced by alanine (K272A) gives monoclinic crystals of space group P2(1) with cell dimensions a = 67.1 A, b = 108.6 A, c = 124.5 A and beta = 115.7 degrees. These crystal forms diffract to 2.4 A and 2.7 A resolution, respectively.

Crystallization↗

PubMed1994

Crystallization of seed globulins from legumes.

Seeds contain large quantities of proteins and are therefore main food sources. In the last century, protein extracts of legume seeds were dialysed against distilled water and in some cases small crystals of pure protein appeared. However, those crystals were generally of poor quality with respect to X-ray diffraction. Recently, the crystallization of some of them was improved and the structures of two 7S globulins, phaseolin from Phaseolus vulgaris and canavalin from Canavalia ensiformis, have been determined at 3.0 and 2.6 A resolution, respectively. Efforts to improve the quality of the phaseolin crystals resulted in three new crystal forms which will be discussed in this paper. The only high-resolution X-ray analysis of a seed globulin from legumes is that of narbonin, a 2S protein from Vicia narbonensis. The crystal structure at 1.8 A shows a very compact packing in layers of molecules. The intermolecular contacts include salt bridges and hydrophobic clusters that might facilitate both the aggregation of the molecules and their crystallization. Because the seed globulins appear in large quantities in the protein bodies of the seeds, efficient packing of the molecules similar to the crystal packing can be assumed.

Journal Article↗

PubMed1993

A monoclonal antibody Fab fragment crystallized with and without a peptide epitope from HIV.

The Fab fragment of CB 4-1, a monoclonal murine antibody against HIV protein p24, has been produced. It forms a complex with a synthetic antigen, an epitope of p24 made up of 11 amino acids, with the binding constant Kd = 3.6 x 10(-9) M. Crystals of hexagonal and orthorhombic space group has been obtained by cocrystallization of the Fab with the epitope and crystallization without the epitope, respectively. In either case, the crystals are suitable for X-ray structural analysis. Crystals of the Fab fragment cocrystallized with the peptide have the space group P 6(3)22 with cell dimensions of a = b = 105 A, c = 297 A. Fab crystals without the epitope are in space group C 222 with cell dimensions a = 110.1 A, b = 110.2 A, c = 150.1 A.

Animals↗

PubMed1993

A double-blind comparison of the effects of carvedilol and captopril on serum lipid concentrations in patients with mild to moderate essential hypertension and dyslipidaemia.

We have studied 250 patients with mild to moderate essential hypertension (diastolic blood pressure 95-114 mmHg) and dyslipidaemia (high-density lipoprotein cholesterol (HDL-C) below 1.03 mmol.l-1, total cholesterol 5.17-9.05 mmol.l-1, and triglycerides 2.26-5.64 mmol.l-1) in a controlled double-blind, multicentre, parallel group trial. The patients took a fat-modified diet. After a 4-week placebo period, patients who continued to fulfil the selection criteria were randomly allocated to treatment with either carvedilol (a vasodilating beta-blocker) 25-50 mg o.d. (n = 116) or captopril (an ACE inhibitor) 25-50 mg o.d. (n = 117) for 6 months. In both groups there were favourable effects on the serum lipids. The relative changes (medians) in the carvedilol and captopril group were respectively: increase in HDL-C by 11% and 8%, decrease in total cholesterol by 11% and 10%, in low-density lipoprotein cholesterol by 16% and 12%, and in triglycerides by 13% and 14%. Equivalence of the two treatments was confirmed for the target variable change in HDL-C at a significance level of 5%. Reductions in supine systolic/diastolic blood pressures were comparable in the two groups (carvedilol: 23/19 mmHg, captopril: 20/18 mmHg). The improvement in lipid metabolism in patients treated with carvedilol is probably due to its alpha 1-blocking properties.

Adult↗

PubMed1992

A TIM barrel protein without enzymatic activity? Crystal-structure of narbonin at 1.8 A resolution.

The major protein component in seeds is storage protein. These have no known enzymatic activity and act to provide amino acids as a source of metabolites in the developing seedling. We report here the first three dimensional crystal structure of a seed storage globulin at high resolution. The molecule of the 2S globulin, narbonin, from Vicia narbonensis L., consists of an eight-stranded parallel alpha/beta barrel structure similar to that observed in triose phosphate isomerase (TIM). Narbonin is the first protein with this topology possessing no known enzymatic activity. Because of the lack of sequence information most of the primary structure was determined directly from the electron density.

Amino Acid Sequence↗

PubMed1990

Narbonin, a 2 S globulin from Vicia narbonensis L. Crystallization and preliminary crystallographic data.

A seed globulin from Vicia narbonensis L. has been crystallized by vapour diffusion induced pH-shift. Crystals are suitable for high-resolution X-ray structural analysis and diffract to better than 1.5 A. Narbonin crystallizes in the monoclinic space group P21 with alpha = 46.9 A, b = 75.5 A, c = 50.9 A, alpha = gamma = 90 degrees, beta = 120.5 degrees. The protein consists of one polypeptide chain that does not coincide with the subunits of legumin or vicilin after SDS/polyacrylamide gel electrophoresis and has a relative molecular mass of about 33,000.

Crystallization↗

PubMed1990

Blood-pressure-lowering effect of carvedilol vs nitrendipine in geriatric hypertensives.

Carvedilol and nitrendipine were given for 12 weeks in a double-blind study to 81 elderly patients (greater than or equal to 60 years) with essential hypertension. The effects on blood pressure were measured (Riva Rocci) before medication and after 2 h with the patient in a lying and standing position after 4 weeks of placebo therapy as well as after 4, 8 and 12 weeks of treatment. Carvedilol (25 mg/o.d.) reduced blood pressure measured in the supine and erect position very successfully, similar to the reduction achieved with nitrendipine (20 mg/o.d.), without influencing the pulse rate. Both substances were well tolerated. Carvedilol is an alternative substance for lowering high blood pressure in elderly hypertensive patients.

Adrenergic beta-Antagonists↗

PubMed1970