The base catalysed anomerisation of β-5-formyluridine; crystal and molecular structure of α-5-formyluridine

On treatment with strong base β-5-formyluridine undergoes an anomerisation to give a mixture of the α- and β-anomers. The anomers have been separated by fractional recrystallisation and the absolute configuration of the α-anomer has been determined by X-ray analysis.     

The crystal and molecular structure of α-laminaribiose octa-acetate

The crystal and molecular structure of octa-O-acetyl-α-laminaribiose is compared with those of disaccharides, and their acetylated derivatives, related to polysaccharides of the laminarin and curdlan type. Marked differences are found in the endo- and exo-cyclic torsion angles as well as in the molecular geometry of the glycosidic linkage. The conformation of AcO-6′ provides the first experimental evidence of the theoretically predicted gauche-trans-gauche⁻ conformation. This conformational change, together with the α-orientation of AcO-1, alters the overall shape of the molecule and influences the packing features. The effect of the acetylation is also examined in terms of calculated conformational energy maps.     

Evidence for the occurrence of two forms of β-lipotropin in rat pituitary

A peptide isolated from porcine gut according to its glucagon-like activity in liver (bioactive enteroglucagon) has been characterized immunologically, biologically and chemically: its potency relative to pancreatic glucagon in interacting with an antiglucagon antibody, hepatic glucagon-binding sites and hepatic adenylate cyclase was ～100%, 20% and 10%, respectively. In contrast, it is ～20-times more potent than glucagon in oxyntic glands, justifying the term ‘oxyntomodulin’. Chemically, it consists in the 29 amino acid-peptide glucagon elongated at its C-terminal end by the octapeptide Lys—Arg—Asn—Lys—Asn—Asn—Ile &;—Ala; accordingly, it is called ‘glucagon-37’     

The effects of cholesterol and β-sitosterol on the structure of saturated diacylphosphatidylcholine bilayers

The structures of DMPC and DPPC bilayers in unilamellar liposomes, in the presence of 33.3 mol% cholesterol or the plant sterol β-sitosterol, have been studied by small-angle neutron scattering. The bilayer thickness d _L increases in a similar way for both sterols. The repeat distance in multilamellar liposomes, as determined by small-angle X-ray diffraction, is larger in the presence of β-sitosterol than in the presence of cholesterol. We observe that each sterol modifies the interlamellar water layer differently, cholesterol reducing its thickness more efficiently than β-sitosterol, and conclude that cholesterol suppresses bilayer undulations more effectively than β-sitosterol.     

Optimizing the crystal size and habit of β-sitosterol in suspension

The aim of this work was to survey how processing parameters affect the crystal growth of β-sitosterol in suspension. The process variables studied were the cooling temperature, stirring time and stirring rate during recrystallization. In addition, we investigated the effect a commonly used surfactant, polysorbate 80, has on crystal size distribution and the polymorphic form. This study describes the optimization of the crystallization process, with the object of preparing crystals as small as possible. Particle size distribution and habit were analyzed using optical microscopy, and the crystal structure was analyzed using X-ray diffractometry. The cooling temperature had a remarkable influence on the crystal size. Crystals with a median crystal length of ≈23 μm were achieved with a low cooling temperature (<10°C); however, a fairly large number of crystals over 50 μm appeared. Higher cooling temperatures (>30°C) caused notable crystal growth both in length and width. Rapid (250 rpm), continuous stirring until the suspensions had cooled to room temperature created small, less than 50 μm long (median <20 μm), needle-shaped crystals. The addition of surfactant slightly reduced the size of the initially large crystals. Both hemihydrate and monohydrate crystal forms occurred throughout, regardless of the processing parameters. By using an optimized process, it was possible to obtain a microcrystalline suspension, with a smooth texture.     

Molecular nature of β-Galactosidase from different tissues in two strains of the house mouse

One inbred mouse strain, C57BL/Kl, has high galactosidase activities in all tissues while another strain, DBA/2/Kl, has low activities determined by the Bgs locus. Beta-Galactosidase from these two strains was partly purified by a five-step procedure: acidification, ammonium sulfate precipitation, gel filtration at two pHs, and isoelectric focusing. No qualitative differences were found between the enzyme preparations from the two strains. They had identical heat inactivation curves, pH optima, molecular weight, and isoelectric points, and the Km values were very similar. It thus seems that this genetic difference in enzyme activity probably cannot be explained by a variation of the galactosidase-specific activity but rather reflects a difference in number of enzyme molecules. Eight different isoenzymes were separated from liver, kidney, and spleen. Each isoenzyme has a different electrophoretic mobility and there is a stepwise increase in molecular weight from 143,000 to 380,000 beginning with the protein having the lowest isoelectric point. A likely interpretation is that the isoenzymes bind a smaller polypeptide in varying numbers in addition to the enzymatic polypeptide per se.     

The effect of quaternary structure on the state of the α and β subunits within nitrosyl haemoglobin: Low temperature photodissociation and the ESR spectra

Photodissociation of nitrosyl haemoglobin and nitrosyl hybrids, in which either the α or β subunit is in the nitrosyl form has been studied at liquid helium temperature (4.2°K) by electron spin resonance and optical absorption spectroscopy. In the presence of inositol hexaphosphate, the photodissociated form of nitrosyl haemoglobin showed an anomalous absorption spectrum in the near infrared region. The experiments with nitrosyl hybrids showed that the α^NO subunit within the T state haemoglobin is predominantly responsible for the anomalous photodissociated form and the ESR spectrum with three distinct hyperfines. The ESR spectrum of α₂^NOβ₂^deoxy with inositol hexaphosphate appeared to be very similar to that of the 5-coordinated NO-haem complexes but the absorption spectrum of its photodissociated form was similar to none of protoporphyrin Fe(II) derivatives so far reported. This result suggests that the anomalous photodissociated form may be attributable to some structural distortion of porphyrin or a new electronic state of the haem with different spin state from that of deoxyhaemoglobin.     

Circular dichroism spectra of β-peptides: sensitivity to molecular structure and effects of motional averaging

Circular dichroism spectra of two -peptides, i.e. peptides composed of -amino acids, calculated using ensembles of configurations obtained by molecular dynamics simulation are presented. The calculations were based on 200 ns simulations of a -heptapeptide in methanol at 298 K and 340 K and a 50 ns simulation of a -hexapeptide in methanol at 340 K. In the simulations the peptides sampled both folded (helical) and unfolded structures. Trajectory structures with common backbone conformations were identified and grouped into clusters. The CD spectra were calculated for individual structures, based on peptide-group dipole transition moments obtained from semi-empirical molecular orbital theory and using the so-called matrix method. The single-structure spectra were then averaged over entire trajectories and over clusters of structures. Although certain features of the experimental CD spectra of the -peptides are reproduced by the trajectory-average spectra, there exist clear differences between the two sets of spectra in both wavelength and peak intensities. The analysis of individual contributions to the average spectra shows that, in general, the interpretation of a CD signal in terms of a single structure is not possible. Moreover, there is a large variation in the CD spectra calculated for a set of individual structures that belong to the same cluster, even when a structurally tight clustering criterion is used. This indicates that the CD spectra of these peptides are very sensitive to small local structural differences.     

Three-dimensional structure of deoxycholate-treated purple membrane at 6 Å resolution and molecular averaging of three crystal forms of bacteriorhodopsin

The three-dimensional structure of the deoxycholate-treated form of purple membrane has been determined to a resolution of about 6 Å. Using low temperature electron diffraction data, room temperature electron microscope images and improved methods of data analysis, higher resolution has been reached than was obtained using native membranes of the same size. Statistical analysis of the data shows that the new map is considerably better than earlier maps. The map indicates the probable sites for the lipid molecules that remain in the deoxycholate-treated membranes; some of these sites differ from those suggested by the projection map of Glaeser et al. (1985). Comparison of the bacteriorhodopsin structures now determined independently from three crystal forms shows that the monomer structure is independent of the detailed contacts with lipid molecules. The average of the three structures gives a picture with very little noise showing seven similar rod-like features which are clearly best interpreted as -helices; there is no indication that part of the structure is -sheet as suggested by Jap et al. (1983). Phases from the averaged structure at 6 Å resolution will enable better refinement of the parameters that will be required in the analysis of higher resolution images from tilted specimens needed to extend the projection map at 3.5 Å resolution (Henderson et al. 1986) to produce a three-dimensional atomic resolution map.     

Molecular and crystal structure of the regenerated form of (1→3)-α-d-glucan

The crystal structure of a regenerated form of (1→3)-α-d-glucan, obtained by solid state deacetylation of the triacetate derivative, has been determined by combined X-ray diffraction analysis and stereochemical model refinement. The structure crystallizes in an orthorhombic unit cell with parameters $\text{a = 16.46}\text{A}\text{?}\text{, b = 9.55}\text{A}\text{?}$ and c (fibre repeat)=8.44 Å, and space group P2₁2₁2₁. The chain conformation is nearly completely extended and is very close to a 2/1 helix, even though the dimer residue is the crystallographic repeat unit. An intramolecular O(2)  O(4)′ hydrogen bond stabilizes the conformation and extensive intermolecular hydrogen-bonding abilizes the packing. The resulting structure is sheet-like, with an alternating polarity of chain directions within the sheet. In its sheet-like character, extensive hydrogen-bonding, and insolubility in water, this polymorph of (1→3)-α-d-glucan resembles regenerated cellulose. The reliability of the structure analysis is indicated by the X-ray residual R=0.206.     

The NMR solution structure and characterization of pH dependent chemical shifts of the β-elicitin, cryptogein

The NMR structure of the 98 residue -elicitin, cryptogein, which induces a defence response in tobacco, was determined using 15N and 13C/15N labelled protein samples. In aqueous solution conditions in the millimolar range, the protein forms a discrete homodimer where the N-terminal helices of each monomer form an interface. The structure was calculated with 1047 intrasubunit and 40 intersubunit NOE derived distance constraints and 236 dihedral angle constraints for each subunit using the molecular dynamics program DYANA. The twenty best conformers were energy-minimized in OPAL to give a root-mean-square deviation to the mean structure of 0.82 Å for the backbone atoms and 1.03 Å for all heavy atoms. The monomeric structure is nearly identical to the recently derived X-ray crystal structure (backbone rmsd 0.86 Å for residues 2 to 97) and shows five helices, a two stranded antiparallel -sheet and an -loop. Using 1H,15N HSQC spectroscopy the pKa of the N- and C-termini, Tyr12, Asp21, Asp30, Asp72, and Tyr85 were determined and support the proposal of several stabilizing ionic interactions including a salt bridge between Asp21 and Lys62. The hydroxyl hydrogens of Tyr33 and Ser78 are clearly observed indicating that these residues are buried and hydrogen bonded. Two other tyrosines, Tyr47 and Tyr87, show pKa's >12, however, there is no indication that their hydroxyls are hydrogen bonded. Calculations of theoretical pKa's show general agreement with the experimentally determined values and are similar for both the crystal and solution structures.     

The localization and the isoenzymes of α- and β-Galactosidases in root tips

The localization was studied of α- and β-galactosidases in frozen sections of Ca-formol fixed root tips using simultaneous azocoupling reaction. In all species studied (Allium cepa,Cucurbita maxima, Lupinus albus, Pisum sativum, Vicia faba, Zea mays) positive results were obtained, the localization being ubiquitous (according to localization typology given here). InVicia faba andZea mays the isoenzymes of α- and β-galactosidases were revealed by means of acrylamide gel electrophoresis, using authors’ modification of Reisfeld method, in whole root tips, particular growth zones and separately in cortex and central cylinder. No differences were observed comparing stele and cortex. Whereas characteristic isoenzyme patterns were found in individual growth zones in maize, no differences appeared in broad bean. A comparison was made of thein situ localization and of the isoenzyme patterns of α- and β-galactosidases with α- and β-glucosidases. In the case of galactosidases, positive results appear with both α- and β-galactoside. The rising of pH to neutrality leads to considerable decrease in the activity of both galactosidases.