Enzymic synthesis,chemical characterisation and Sda activity of GalNAcß1-4[NeuAcα2-3]Galß1-4GlcNAc and GalNAcß1-4[NeuAcα2-3]Galß1-4Glc

The tetrasaccharides GalNAcß1-4[NeuAc2-3]Galß1-4Glc and GalNAcß1-4[NeuAc2-3]Galß1-4GlcNAc were synthesised by enzymic transfer of GalNAc from UDP-GalNAc to 3-sialyllactose (NeuAc2-3Galß1-4Glc) and 3-sialyl-N-acetyllactosamine (NeuAc2-3Galß1-4GlcNAc). The structures of the products were established by methylation and¹H-500 MHz NMR spectroscopy. In Sd^a serological tests the product formed with 3-sialyl-N-acetyllactosamine was highly active whereas that formed with 3-sialyllactose had only weak activity.     

Sequence dependence of DNA conformational flexibility

By using conformational free energy calculations, we have studied the sequence dependence of flexibility and its anisotropy along various conformational variables of DNA base pairs. The results show the AT base step to be very flexible along the twist coordinate. On the other hand, homonucleotide steps, GG(CC) and AA(TT), are among the most rigid sequences. For the roll motion that would correspond to a bend, the TA step is most flexible, while the GG(CC) step is least flexible. The flexibility of roll is quite anisotropic; the ratio of fluctuations toward the major and minor grooves is the largest for the GC step and the smallest for the AA(TT) and CG steps. Propeller twisting of base pairs is quite flexible, especially of A.T base pairs; propeller twist can reach 19 degrees by thermal fluctuation. We discuss the effect of electrostatic parameters, comparison with available experimental results, and biological relevance of these results.     

Distinct patterns in homooligomer tract sequence context in prokaryotic and eukaryotic DNA

The distributions of the junction sequences of homooligomer tracts of various lengths have been examined in prokaryotic DNA sequences and compared with those of eukaryotes. The general trends in the nearest and next to nearest neighbors to the tracts are similar for both groups. In both prokaryotes and eukaryotes A/T runs are preferentially flanked on either the 5' or the 3' ends by A and/or T. G/C runs are preferentially flanked by G and/or C. There is discrimination against A/T runs flanked by G or C and G/C runs flanked by A or T. However, whereas the distribution of prokaryotic homooligomer tract junction sequences was quite homogeneous, large variations were observed in the 5-fold larger eukaryotic database, increasing in magnitude from tracts of length 2 to 3 to 4 base pairs long. Possible DNA conformational implications and in particular DNA curvature and packaging aspects of prokaryotes and eukaryotes are discussed.     

Approaches to breeding for salinity tolerance - a case study on Porteresia coarctata

Cereals are the world's major source of food for human nutrition. Among these, rice (Oryza sativa) is the most prominent and represents the staple diet for more than two-fifths (2.4 billion) of the world's population, making it the most important food crop of the developing world (Anon., 2000a). Rice production in vast stretches of coastal areas is hampered due to high soil salinity. This is because rice is a glycophyte and it does not grow well under saline conditions. In order to increase rice production in these areas there is a need to develop rice varieties suited to saline environments. Research has shown that Porteresia coarctata, a highly salt tolerant wild relative of rice growing in estuarine soils, is an important material for transferring salt tolerant characteristics to rice. It is quite possible that Porteresia may be used as a parent for evolving better and truly salt resistant varieties. The inadequate results and the difficulties associated with conventional breeding techniques necessitate the use of the tools of crop biotechnology in unravelling some of the characteristics of Porteresia that have been highlighted in this report. In view of the limited resources available for increasing salinity tolerance to the breeders to wild rice germplasm, Porteresia is undoubtedly one of the key source species for elevating salinity tolerance in cultivated rice.     

Equilibrium folding and unfolding pathways for a model protein

The folding–unfolding process of reduced bovine pancreatic trypsin inhibitor was investigated with an idealized model employing approximate free energies. The protein is regarded to consist of only C^α and C^β atoms. The backbone dihedral angles are the only conformational variables and are permitted to take discrete values at every 10°. Intraresidue energies consist of two terms: an empirical part taken from the observed frequency distributions of (?,ψ) and an additional favorable energy assigned to the native conformation of each residue. Interresidue interactions are simplified by assuming that there is an attractive energy operative only between residue pairs in close contact in the native structure. A total of 230,000 molecular conformations, with no atomic overlaps, ranging from the native state to the denatured state, are randomly generated by changing the sampling bias. Each conformation is classified according to its conformational energy, F; a conformational entropy, S(F) is estimated for each value of F from the number of samples. The dependence of S(F) on energy reveals that the folding–unfolding transition for this idealized model is an “all-or-none” type; this is attributable to the specific long-range interactions. Interresidue contact probabilities, averaged over samples representing various stages of folding, serve to characterize folding intermediates. Most probable equilibrium pathways for the folding–unfolding transition are constructed by connecting conformationally similar intermediates. The specific details obtained for bovine pancreatic trypsin inhibitor are as follows: (1) Folding begins with the appearance of nativelike medium-range contacts at a β-turn and at the α-helix. (2) These grow to include the native pair of interacting β-strands. This state includes intact regular secondary conformations, as well as the interstrand sheet contacts, and corresponds to an activated state with the highest free energy on the pathway. (3) Additional native long-range contacts are completely formed either toward the amino terminus or toward the carboxyl terminus. (4) In a final step, the missing contacts appear. Although these folding pathways for this model are not consistent with experimental reports, it does indicate multiple folding pathways. The method is general and can be applied to any set of calculated conformational energies and furthermore permits investigation of gross folding features.     

Conformations of folded proteins in restricted spaces

A new method is presented to examine the complete range of folded topologies accessible in the compact state of globular proteins. The procedure is to generate all conformations, with volume exclusion, upon a lattice in a space restricted to the individual protein's known compact conformational space. Using one lattice point per residue, we find 10(2)-10(4) possible compact conformations for the five small globular proteins studied. Subsequently, these conformations are evaluated in terms of residue-specific, pairwise contact energies that favor nonbonded, hydrophobic interactions. Native structures for the five proteins are always found within the best 2% of all conformers generated. This novel method is simple and general and can be used to determine a small group of most favorable overall arrangements for the folding of specific amino acid sequences within a restricted space.     

Solvent effect on binding thermodynamics of biopolymers

The indirect solvent-induced effect on the free energy of binding of biopolymers is examined within the framework of classical statistical mechanics. We focus specifically on the role of the solute-solvent hydrogen bonding. In particular, we have estimated the first order solvent effect on the indirect interaction between two biopolymers. We find that the solvent-induced interactions between two hydrophilic groups through water-bridged hydrogen bonds could significantly enhance the binding free energy. Some preliminary estimates indicate that this effect is significant and perhaps could be crucial in molecular recognition processes. Furthermore, we have calculated, from crystal structure data, the distance distribution between all the oxygens and nitrogens on the surface of some proteins that do not belong to the binding domain. In most cases we found an enhanced peak in the range of 4-5 A, which is where we expect to find strong solvent-induced interactions.     

Effect of neurotensin on pancreatic function in man

The effect of neurotensin on submaximally-stimulated hepatobiliary and pancreatic secretion was studied in 6 healthy subjects. An intravenous infusion of neurotensin 1.4 ± 0.3 pmol/kg/min, designed to reproduce plasma neurotensin immunoreactivity levels within the physiological range, produced a significant increase in pancreatic bicarbonate output. Plasma concentrations of pancreatic polypeptide rose by 83 ± 16 pmol/l and were associated with a small reduction in trypsin, but no significant change in bilirubin outputs.