Computation techniques in the conformational analysis of carbohydrates

A growing number of modern studies of carbohydrates is devoted to spatial mechanisms of their participation in the cell recognition processes and directed design of inhibitors of these processes. No progress in this field is possible without the development of theoretical conformational analysis of carbohydrates. In this review, we generalize literature data on the potentialities of using various molecular-mechanic force fields, the methods of quantum mechanics, and molecular dynamics to study the conformation of glycoside linkage. A possibility of analyzing the reactivity of carbohydrates with the computation techniques is also discussed in brief.     

Absence of Gup1p in Saccharomyces cerevisiae results in defective cell wall composition, assembly, stability and morphology

Saccharomyces cerevisiae Gup1p and its homologue Gup2p, members of the superfamily of membrane-bound O-acyl transferases, were previously associated with glycerol-mediated salt-stress recovery and glycerol symporter activity. Several other phenotypes suggested Gup1p involvement in processes connected with cell structure organization and biogenesis. The gup1Delta mutant is also thermosensitive and exhibits an altered plasma membrane lipid composition. The present work shows that the thermosensitivity is independent of glycerol production and retention. Furthermore, the mutant grows poorly on salt, ethanol and weak carboxylic acids, suggestive of a malfunctioning membrane potential. Additionally, gup1Delta is sensitive to cell wall-perturbing agents, such as Calcofluor white, Zymolyase, lyticase and sodium dodecyl sulphate and exhibits a sedimentation/aggregation phenotype. Quantitative analysis of cell wall components yielded increased contents of chitin and beta-1,3-glucans and lower amounts of mannoproteins. Consistently, scanning electron microscopy showed a strikingly rough surface morphology of the mutant cells. These results suggest that the gup1Delta is affected in cell wall assembly and stability, although the Slt2p/MAP kinase from the PKC pathway was phosphorylated during hypo-osmotic shock to a normal extent. Results emphasize the pleiotropic nature of gup1Delta, and are consistent with a role of Gulp1p in connection with several pathways for cell maintenance and construction/remodelling.     

Disruption of Cortical Microtubules by Overexpression of Green Fluorescent Protein-Tagged α-Tubulin 6 Causes a Marked Reduction in Cell Wall Synthesis

It has been known that the transverse orientation of cortical microtubules （MTs） along the elongation axis is essential for normal cell morphogenesis, but whether cortical MTs are essential for normal cell wall synthesis is still not clear. In the present study, we have investigated whether cortical MTs affect cell wall synthesis by direct alteration of the cortical MT organization in Arabidopsis thaliana. Disruption of the cortical MT organization by expression of an excess amount of green fluorescent protein-tagged a-tubulin 6 （GFP-TUA6） in transgenic Arabidopsis plants was found to cause a marked reduction in cell wall thickness and a de- crease in the cell wall sugars glucose and xylose. Concomitantly, the stem strength of the GFP-TUA6 overexpressors was markedly reduced compared with the wild type. In addition, expression of excess GFP- TUA6 results in an alteration in cell morphogenesis and a severe effect on plant growth and development. Together, these results suggest that the proper organization of cortical MTs is essential for the normal synthesis of plant cell walls.     

Cell elongation in Arabidopsis hypocotyls involves dynamic changes in cell wall thickness

Field-emission scanning electron microscopy was used to measure wall thicknesses of different cell types in freeze-fractured hypocotyls of Arabidopsis thaliana. Measurements of uronic acid content, wall mass, and wall volume suggest that cell wall biosynthesis in this organ does not always keep pace with, and is not always tightly coupled to, elongation. In light-grown hypocotyls, walls thicken, maintain a constant thickness, or become thinner during elongation, depending upon the cell type and the stage of growth. In light-grown hypocotyls, exogenous gibberellic acid represses the extent of thickening and promotes cell elongation by both wall thinning and increased anisotropy during the early stages of hypocotyl elongation, and by increased wall deposition in the latter stages. Dark-grown hypocotyls, in the 48 h period between cold imbibition and seedling emergence, deposit very thick walls that subsequently thin in a narrow developmental window as the hypocotyl rapidly elongates. The rate of wall deposition is then maintained and keeps pace with cell elongation. The outer epidermal wall is always the thickest ( approximately 1 mum) whereas the thinnest walls, about 50 nm, are found in inner cell layers. It is concluded that control of wall thickness in different cell types is tightly regulated during hypocotyl development, and that wall deposition and cell elongation are not invariably coupled.     

A scale of metal ion binding strengths correlating with ionic charge, Pauling electronegativity, toxicity, and other physiological effects

Equilibrium constants for binding to plant plasma membranes have been reported for several metal ions, based upon adsorption studies and zeta-potential measurements. LogK values for the ions are these: Al(3+), 4.30; La(3+), 3.34; Cu(2+), 2.60; Ca(2+) and Mg(2+), 1.48; Na(+) and K(+), 0 M(-1). These values correlate well with logK values for ion binding to many organic and inorganic ligands. LogK values for metal ion binding to 12 ligands were normalized and averaged to produce a scale for the binding of 49 ions. The scale correlates well with the values presented above (R(2)=0.998) and with ion binding to cell walls and other biomass. The scale is closely related to the charge (Z) and Pauling electronegativity (PE) of 48 ions (all but Hg(2+)); R(2)=0.969 for the equation (Scale values)=-1.68+Z(1.22+0.444PE). Minimum rhizotoxicity of metal ions appears to be determined by binding strengths: log a(PM,M)=1.60-2.41exp[0.238(Scale values)] determines the value of ion activities at the plasma membrane surface (a(PM,M)) that will ensure inhibition of root elongation. Additional toxicity appears to be related to softness, accounting for the great toxicity of Ag(+), for example. These binding-strength values correlate with additional physiological effects and are suitable for the computation of cell-surface electrical potentials.     

Sentinels at the wall: cell wall receptors and sensors

The emerging view of the plant cell wall is of a dynamic and responsive structure that exists as part of a continuum with the plasma membrane and cytoskeleton. This continuum must be responsive and adaptable to normal processes of growth as well as to stresses such as wounding, attack from pathogens and mechanical stimuli. Cell expansion involving wall loosening, deposition of new materials, and subsequent rigidification must be tightly regulated to allow the maintenance of cell wall integrity and co-ordination of development. Similarly, sensing and feedback are necessary for the plant to respond to mechanical stress or pathogen attack. Currently, understanding of the sensing and feedback mechanisms utilized by plants to regulate these processes is limited, although we can learn from yeast, where the signalling pathways have been more clearly defined. Plant cell walls possess a unique and complicated structure, but it is the protein components of the wall that are likely to play a crucial role at the forefront of perception, and these are likely to include a variety of sensor and receptor systems. Recent plant research has yielded a number of interesting candidates for cell wall sensors and receptors, and we are beginning to understand the role that they may play in this crucial aspect of plant biology.     

Quantitative evaluation of adhesion of lactobacilli isolated from human intestinal tissues to human colonic mucin using surface plasmon resonance (BIACORE assay)

AIMS: To isolate lactobacilli from the mucus layer of the human intestine and evaluate their adhesion abilities using a BIACORE assay. METHODS AND RESULTS: Thirty strains of lactobacilli were isolated from the mucus layer of normal human intestinal tissues using conventional plate culture. The strains were identified using homology comparisons of the 16S rDNA sequence to databases as Lactobacillus salivarius (26%), Lactobacillus fermentum (13%), Lactobacillus gasseri (10%), Lactobacillus paracasei (7%), Lactobacillus casei (3%), Lactobacillus mucosae (3%) and Lactobacillus plantarum (3%). Lactobacillus plantarum LA 318 shows the highest adhesion to human colonic mucin (HCM) using the BIACORE assay at 115.30 +/- 12.37 resonance unit (RU). The adhesion of cell wall surface proteins from strain LA 318 was significantly higher to HCM than to bovine serum albumin (BSA; P < 0.05). CONCLUSIONS: We isolated 30 strains of lactobacilli. Lactobacillus salivarius was the predominant species of lactobacilli isolated in this study. The adhesion of strain LA 318 isolated from human transverse colon to its mucin was shown. The adhesion could be mediated by lectin-like components on the bacterial cell surface. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study where lactobacilli were isolated from human intestinal tissues and shown to adhere to HCM.     

Correlation between cell wall extensibility and the content of diferulic and ferulic acids in cell walls of Oryza sativa coleoptiles grown under water and in air

Rice ( Oryza sativa L. cv. Sasanishiki) coleoptiles grown under water achieved greater length than those grown either in air or under water with constant air bubbling. The extensibility of cell walls in coleoptiles grown under water was larger than that in the other treatments. Per unit length of the coleoptile, the content of ferulic and diferulic acids ester-linked to hemicelluloses was higher in air and bubbling type coleoptiles than in water type ones. The extensibility of the coleoptile cell walls correlated with the content of diferulic acids per unit length and per hemicellulose, suggesting that the enhancement of the formation of diferulic acid bridges in hemicelluloses in air or under water with air bubbling makes the cell walls mechanically rigid; thereby inhibiting cell elongation in rice coleoptiles. In addition, the ratio of diferulic acid to ferulic acid was almost constant irrespective of coleoptile age, zone and growth conditions, suggesting that the feruloylation of hemicelluloses is rate-limiting in the formation of diferulic acid bridges in the cell walls of rice coleoptiles.     

Three-dimensional macro-scale assessment of regional and temporal wall shear stress characteristics on aortic valve leaflets

The aortic valve (AV) achieves unidirectional blood flow between the left ventricle and the aorta. Although hemodynamic stresses have been shown to regulate valvular biology, the native wall shear stress (WSS) experienced by AV leaflets remains largely unknown. The objective of this study was to quantify computationally the macro-scale leaflet WSS environment using fluid–structure interaction modeling. An arbitrary Lagrangian–Eulerian approach was implemented to predict valvular flow and leaflet dynamics in a three-dimensional AV geometry subjected to physiologic transvalvular pressure. Local WSS characteristics were quantified in terms of temporal shear magnitude (TSM), oscillatory shear index (OSI) and temporal shear gradient (TSG). The dominant radial WSS predicted on the leaflets exhibited high amplitude and unidirectionality on the ventricularis (TSM>7.50 dyn/cm², OSI < 0.17, TSG>325.54 dyn/cm² s) but low amplitude and bidirectionality on the fibrosa (TSM < 2.73 dyn/cm², OSI>0.38, TSG < 191.17 dyn/cm² s). The radial WSS component computed in the leaflet base, belly and tip demonstrated strong regional variability (ventricularis TSM: 7.50–22.32 dyn/cm², fibrosa TSM: 1.26–2.73 dyn/cm²). While the circumferential WSS exhibited similar spatially dependent magnitude (ventricularis TSM: 1.41–3.40 dyn/cm², fibrosa TSM: 0.42–0.76 dyn/cm²) and side-specific amplitude (ventricularis TSG: 101.73–184.43 dyn/cm² s, fibrosa TSG: 41.92–54.10 dyn/cm² s), its temporal variations were consistently bidirectional (OSI>0.25). This study provides new insights into the role played by leaflet–blood flow interactions in valvular function and critical hemodynamic stress data for the assessment of the hemodynamic theory of AV disease.