Inferring protein interactions from phylogenetic distance matrices

Finding the interacting pairs of proteins between two different protein families whose members are known to interact is an important problem in molecular biology. We developed and tested an algorithm that finds optimal matches between two families of proteins by comparing their distance matrices. A distance matrix provides a measure of the sequence similarity of proteins within a family. Since the protein sets of interest may have dozens of proteins each, the use of an efficient approximate solution is necessary. Therefore the approach we have developed consists of a Metropolis Monte Carlo optimization algorithm which explores the search space of possible matches between two distance matrices. We demonstrate that by using this algorithm we are able to accurately match chemokines and chemokine-receptors as well as the tgfbeta family of ligands and their receptors.     

Cloning of a human tRNA isopentenyl transferase

A cDNA of human origin is shown to encode a tRNA isopentenyl transferase (E.C. 2.5.1.8). Expression of the gene in a Saccharomyces cerevisiae mutant lacking the endogenous tRNA isopentenyl transferase MOD5 resulted in functional complementation and reintroduction of isopentenyladenosine into tRNA. The deduced amino acid sequence contains a number of regions conserved in known tRNA isopentenyl transferases. The similarity to the S. cerevisiae MOD5 protein is 53%, and to the Escherichia coli MiaA protein 47%. The human sequence was found to contain a single C2H2 Zn-finger-like motif, which was detected also in the MOD5 protein, and several putative tRNA transferases located by BLAST searches, but not in prokaryotic homologues.     

Characterization of pigment apparatus in winter-green and summer-green leaves of a shade-tolerant plant <Emphasis Type="Italic">Ajuga reptans</Emphasis>

Comparative study was performed to assess the content and proportions of photosynthetic pigments and the violaxanthin cycle (VXC) activity in winter-green and summer-green leaves of bugleweed (Ajuga reptans L.) plants grown in shaded (photosynthetically active radiation, PAR 150 μmol/(m² s)) and sunny (PAR 1200 μmol/(m² s)) habitats in the Botanic Garden of Jagiellonian University (Krakow, Poland). In overwintered and newly formed leaves of shade plants, the content of green and yellow pigments was two times higher than in leaves of sun plants. The shade plants were distinguished by accumulation of β-carotene, while lutein was predominant in leaves of sun plants. Under the action of strong light (2000 μmol/(m²s)), the level of violaxanthin deepoxidation in winter-green leaves of shade and sun plants increased five- to sixfold, whereas it changed insignificantly in summer-green leaves of shade plants. It is concluded that, in a shadetolerant species A. reptans, the photosynthetic apparatus of winter-green leaves in sun and shade plants and of summer-green leaves in sun plants is protected against excess insolation by high activity of VXC. The carotenoids of summer-green leaves in shade plants are supposed to function mainly as light-harvesting pigments.     

Comparative study of the "membrane reserve" of blood cells of reptiles and mammals

Based on the method of hypoosmotic loads, the value of membrane reserve and its use by blood cells of reptiles and mammals have been studied. It has been shown that lymphocytes of the both animal species have the highest reserve of plasmalemma, while frog heterophils - the lowest one. A significantly lower part of the membrane reserve participates in formation of phagosomes by mammalian neutrophils as compared with amphibian erythrocytes.     

A CBS domain-containing pyrophosphatase of Moorella thermoacetica is regulated by adenine nucleotides

CBS (cystathionine beta-synthase) domains are found in proteins from all kingdoms of life, and point mutations in these domains are responsible for a variety of hereditary diseases in humans; however, the functions of CBS domains are not well understood. In the present study, we cloned, expressed in Escherichia coli, and characterized a family II PPase (inorganic pyrophosphatase) from Moorella thermoacetica (mtCBS-PPase) that has a pair of tandem 60-amino-acid CBS domains within its N-terminal domain. Because mtCBS-PPase is a dimer and requires transition metal ions (Co2+ or Mn2+) for activity, it resembles common family II PPases, which lack CBS domains. The mtCBS-PPase, however, has lower activity than common family II PPases, is potently inhibited by ADP and AMP, and is activated up to 1.6-fold by ATP. Inhibition by AMP is competitive, whereas inhibition by ADP and activation by ATP are both of mixed types. The nucleotides are effective at nanomolar (ADP) or micromolar concentrations (AMP and ATP) and appear to compete for the same site on the enzyme. The nucleotide-binding affinities are thus 100-10000-fold higher than for other CBS-domain-containing proteins. Interestingly, genes encoding CBS-PPase occur most frequently in bacteria that have a membrane-bound H+-translocating PPase with a comparable PP(i)-hydrolysing activity. Our results suggest that soluble nucleotide-regulated PPases act as amplifiers of metabolism in bacteria by enhancing or suppressing ATP production and biosynthetic reactions at high and low [ATP]/([AMP]+[ADP]) ratios respectively.     

Mitochondrial lipid abnormality and electron transport chain impairment in mice lacking alpha-synuclein

The presynaptic protein alpha-synuclein, implicated in Parkinson disease (PD), binds phospholipids and has a role in brain fatty acid (FA) metabolism. In mice lacking alpha-synuclein (Snca-/-), total brain steady-state mass of the mitochondria-specific phospholipid, cardiolipin, is reduced 22% and its acyl side chains show a 51% increase in saturated FAs and a 25% reduction in essential n-6, but not n-3, polyunsaturated FAs. Additionally, 23% reduction in phosphatidylglycerol content, the immediate biosynthetic precursor of cardiolipin, was observed without alterations in the content of other brain phospholipids. Consistent with these changes, more ordered lipid head group and acyl chain packing with enhanced rotational motion of diphenylhexatriene (DPH) about its long axis were demonstrated in time-resolved DPH fluorescence lifetime experiments. These abnormalities in mitochondrial membrane properties were associated with a 15% reduction in linked complex I/III activity of the electron transport chain, without reductions in mitochondrial number, complex II/III activity, or individual complex I, II, III, or IV activity. Reduced complex I activity is thought to be a critical factor in the development of PD. Thus, altered membrane composition and structure and impaired complex I/III function in Snca-/- brain suggest a relationship between alpha-synuclein's role in brain lipid metabolism, mitochondrial function, and PD.     

Calcitriol inhibits growth response to Platelet-Derived Growth Factor-BB in human prostate cells

Calcitriol, a hormonal form of Vitamin D, regulates growth of normal and cancer cells of various origins by modulation of peptide growth factors signaling. Platelet-Derived Growth Factor (PDGF) signaling pathway is involved in prostate cancer progression. We studied the expression of PDGF receptors in human prostate primary stromal cells and cancer epithelial cell lines and growth response to PDGF-BB isoform. We found that the expression of PDGF receptors and PDGF-BB-mediated cell growth are regulated by calcitriol in prostate cells. Quantitative RT-PCR analysis revealed a lower level of mRNA for PDGF receptors in LNCaP and PC-3 cells than in primary stromal cells. Western blotting showed a high amount of PDGFRalpha and beta proteins in primary stromal cells that could not be detected in LNCaP, which may explain the resistance of LNCaP cells to growth-promoting effect of PDGF-BB. Addition of Epidermal Growth Factor (EGF) to the culture medium induces the expression of PDGFRbeta and restores responsiveness of LNCaP to PDGF-BB to some extent. Calcitriol down-regulates PDGFRbeta expression and negatively regulates PDGF-mediated cell growth. Calcitriol does not affect PDGFRalpha and PDGF-B mRNA expression. We suggest that inhibition of PDGFRbeta expression by calcitriol might reduce responsiveness of prostate cells to mitogenic action of PDGF-BB.     

Seasonal changes of violaxanthin cycle pigment de-epoxidation in wintergreen and evergreen plants

We studied carotenoids composition and the activities of the xanthophylls pigments in evergreen conifers (Abies sibirica, Juniperus communis, Picea obovata) and dwarf-shrub (Vaccinium vitis-idaea), and in wintergreen herbaceous plants (Ajuga reptans, Pyrola rotundifolia) growing near Syktyvkar (61°67(/) N 50°77(/) E). The carotenoid pool consisted mainly of following xanthophylls: lutein (70%), neoxanthin (7-10%) and a xanthophylls cycle component - violaxanthin (3-15%). Zeaxanthin and antheraxanthin were found in conifer samples collected in December-March while in other species - during all year. A direct connection between xanthophyll pigment de-epoxidation level and light energy thermal dissipation was shown only for boreal conifer species. It is proposed that zeaxanthin plays a central role in the dissipation of excess excitation energy (nonphotochemical quenching) in the antenna of photosystem II (PSII). We conclude that the increase in the extent of de-epoxidation is beneficial for the retention of PSII activity for conifers in early spring and for herbs in summer.     

Phosphoramidate derivatives of acyclovir: Synthesis and antiviral activity in HIV-1 and HSV-1 models in vitro

The antiviral activity against HIV and HSV and the chemical stability of ACV phosphoramidate derivatives were studied. The phosphoramidates of ACV demonstrated moderate activity. The best compound appeared to be 9-(2-hydroxymethyl)guanine phosphoromonomorpholidate (7), which inhibited virus replication in pseudo-HIV-1 particles by 50% at 50 μM. It also inhibited replication of wild-type HSV-1 (9.7 μM) as well as an acyclovir-resistant strain (25 μM). None of the synthesised compounds showed any cytotoxicity.     

Mesostructure and activity of photosynthetic apparatus for three crassulacean species grown in cold climate

A comparative study of leaf anatomy and morphology and of CO₂ exchange was conducted with Rhodiola rosea L., Hylotelephium triphyllum (Haw.) Holub., and Sedum acre L. as representative Crassulacean species occurring in the northeast European Russia. The leaf mesophyll in R. rosea was clearly differentiated into the palisade and spongy tissues, whereas the mesophyll of stonecrops (H. triphyllum and S. acre) was composed of round-shaped cells. The leaves of S. acre featured the largest volume of mesophyll cells and possessed water-retaining cells located around conducting bundles. The chloroplast volume in S. acre (50 μm³) was three times smaller and the number of chloroplasts per cell (170 cell^?1) was three times higher than in R. rosea and H. triphyllum (50–55 cell^?1). The content of chlorophylls (5–7 mg/g dry wt) and carotenoids (1.5–2.0 mg/g dry wt) in R. rosea leaves was 2–3 times higher than in leaves of stonecrops. The rate of CO₂ net uptake in Crassulacean species depended on mesostructure and correlated with the content of pigments and soluble carbohydrates. The photosynthetic rate in R. rosea under optimal irradiance and temperature attained the value of 40 mg/(g dry wt), which is 3 and 8 times higher than in H. triphyllum and S. acre, respectively. The temperature optimum for photosynthesis of R. rosea was observed at 8–18°C, while the optimum for stonecrops was shifted towards higher temperatures by 3–5°C. At chilling temperatures (5–7°C), the leaves of R. rosea retained 50% of their maximal photosynthetic rate, while photosynthetic rates in H. triphyllum and S. acre leaves lowered to 25–30% of the maximal rate. The increase in temperature to 25–30°C led to depression of CO₂ net uptake in leaves of Crassulacean species. In R. rosea and H. triphyllum, the rate of photosynthetic electron flow was depressed at high irradiances and temperatures that were supraoptimal for net photosynthesis. It is concluded that the photosynthetic apparatus of Crassulacean species is well adapted to moderate and chilling temperatures, which adjusts the plant metabolism to “life strategies” under conditions of cold climate.