The HSPGs Syndecan and Dallylike bind the receptor phosphatase LAR and exert distinct effects on synaptic development

The formation and plasticity of synaptic connections rely on regulatory interactions between pre- and postsynaptic cells. We show that the Drosophila heparan sulfate proteoglycans (HSPGs) Syndecan (Sdc) and Dallylike (Dlp) are synaptic proteins necessary to control distinct aspects of synaptic biology. Sdc promotes the growth of presynaptic terminals, whereas Dlp regulates active zone form and function. Both Sdc and Dlp bind at high affinity to the protein tyrosine phosphatase LAR, a conserved receptor that controls both NMJ growth and active zone morphogenesis. These data and double mutant assays showing a requirement of LAR for actions of both HSPGs lead to a model in which presynaptic LAR is under complex control, with Sdc promoting and Dlp inhibiting LAR in order to control synapse morphogenesis and function.     

Solution structure and DNA-binding properties of the phosphoesterase domain of DNA ligase D

The phosphoesterase (PE) domain of the bacterial DNA repair enzyme LigD possesses distinctive manganese-dependent 3′-phosphomonoesterase and 3′-phosphodiesterase activities. PE exemplifies a new family of DNA end-healing enzymes found in all phylogenetic domains. Here, we determined the structure of the PE domain of Pseudomonas aeruginosa LigD (PaePE) using solution NMR methodology. PaePE has a disordered N-terminus and a well-folded core that differs in instructive ways from the crystal structure of a PaePE•Mn²⁺• sulfate complex, especially at the active site that is found to be conformationally dynamic. Chemical shift perturbations in the presence of primer-template duplexes with 3′-deoxynucleotide, 3′-deoxynucleotide 3′-phosphate, or 3′ ribonucleotide termini reveal the surface used by PaePE to bind substrate DNA and suggest a more efficient engagement in the presence of a 3′-ribonucleotide. Spectral perturbations measured in the presence of weakly catalytic (Cd²⁺) and inhibitory (Zn²⁺) metals provide evidence for significant conformational changes at and near the active site, compared to the relatively modest changes elicited by Mn²⁺.     

Status of arbuscular mycorrhizal fungi (AMF) in the Sundarbans of India in relation to tidal inundation and chemical properties of soil

The arbuscular mycorrhizal status of fifteen mangroves and one mangrove associate was investigated from 27 sites of three inundation types namely, diurnal, usual springtide and summer springtide. Roots and rhizospheric soil samples were analysed for spore density, frequency of mycorrhizal colonization and some chemical characteristics of soil. Relative abundance, frequency and spore richness of AMF were assessed at each inundation type. All the plant species except Avicennia alba exhibited mycorrhizal colonization. The study demonstrated that mycorrhizal colonization and spore density were more influenced by host plant species than tidal inundation. Forty four AMF species belonging to six genera, namely Acaulospora, Entrophospora, Gigaspora, Glomus, Sclerocystis and Scutellospora, were recorded. Glomus mosseae exhibited highest frequency at all the inundation types; Glomus fistulosum, Sclerocystis coremioides and Glomus mosseae showed highest relative abundance at sites inundated by usual springtides, summer springtides and diurnal tides, respectively. Spore richness of AMF was of the order usual springtide > diurnal > summer springtide inundated sites. The mean spore richness was 3.27. Diurnally inundated sites had the lowest concentrations of salinity, available phosphorus, exchangeable potassium, sodium and magnesium. Statistical analyses indicated that mycorrhizal frequency and AMF spore richness were significantly negatively correlated to soil salinity. Spore richness was also significantly negatively correlated to available phosphorus. The soil parameters of the usual springtide inundated sites appeared to be favourable for the existence of maximum number of AMF. Glomus mosseae was the predominant species in terms of frequency in the soils of the Sundarbans.     

Photosynthesis and water-use characteristics in Indian mangroves

Photosynthesis and water efflux were measured in different PAR and stomatal conductance in members of Avicenniaceae and Rhizophoraceae. Trend of leaf temperature with irradiance and its effect on photosynthesis were also estimated. In most of the studied species, photosynthesis and stomatal conductance followed similar trends with increase in irradiance. The rate of net photosynthesis and stomatal conductance were higher in members of Avicenniaceae than in Rhizophoraceae. In Avicenniaceae, the optimum PAR for maximum photosynthesis ranged between 1340–1685 (μmol m^-2s^-1, which was also higher than that of Rhizophoraceae (840-1557 μmol m^-2s^-1). Almost in all the studied taxa, transpiration and stomatal conductance followed similar trends and reached the maximal peaks at the same PAR value. The range of breakeven leaf temperature was almost the same in both the families (34-36°C in Avicenniaceae and 33.5-36.3°C in Rhizophoraceae), beyond which assimilation rate declined.     

Conjugation of methotrexate to IgG antibodies and their F(ab)2 fragments and the effect of conjugated methotrexate on tumor growth in vivo

Summary Methotrexate (MTX) was first conjugated to antibovine serum albumin IgG (antiBSA) or its F(ab)₂ fragment to define conditions for retention of drug and antibody activity. With identical drug: protein molar ratios, incorporation in the F(ab)₂ fragment was lower than in intact antiBSA, an observation consistent with analysis of the number of lysine residues (22 in F(ab)₂ compared to 40 in antiBSA). In either case, up to approximately 10 mol MTX could be incorporated per mol protein, with recovery of 70% of the protein. At an incorporation ratio of 6 mol MTX per mol protein, MTX-antiBSA retained 100% of antibody activity and MTX-F(ab)₂antiBSA retained 75%. MTX-antiBSA and MTX-F(ab)₂antiBSA were equally potent in vitro inhibitors of dihydrofolate reductase. Conjugates prepared from antiEL4 IgG (AELG) and from F(ab)₂AELG significantly increased survival in EL4 lymphoma-bearing mice compared with mice receiving equal amounts (5 mg MTX/kg) of free MTX, MTX linked to the F(ab)₂ fragment of normal rabbit IgG, or a simple mixture of MTX and F(ab)₂AELG. MTX-AELG at this dose level produced longer survival than MTX-F(ab)₂AELG (0.0052AELG MTX linked to the F(ab)₂ fragment of AELG - MTX-F(ab)₂antiBSA MTX linked to the F(ab)₂ fragment of antiBSA - MTX-F(ab)₂NRG MTX linked to the F(ab)₂ fragment of NRG - MTX-NRG MTX linked to NRG - NHS N-hydroxysuccinimide - NRG normal rabbit IgG - PBS 0.01 M sodium phosphate (pH 7.1) containing 0.45 M sodium chloride - TAA tumor-associated antigen - t_1/2 half-life     

A novel, specific interaction involving the Csk SH3 domain and its natural ligand.

C-terminal Src kinase (Csk) takes part in a highly specific, high affinity interaction via its Src homology 3 (SH3) domain with the proline-enriched tyrosine phosphatase PEP in hematopoietic cells. The solution structure of the Csk-SH3 domain in complex with a 25-residue peptide from the Pro/Glu/Ser/Thr-rich (PEST) domain of PEP reveals the basis for this specific peptide recognition motif involving an SH3 domain. Three residues, Ala 40, Thr 42 and Lys 43, in the SH3 domain of Csk specifically recognize two hydrophobic residues, Ile 625 and Val 626, in the proline-rich sequence of the PEST domain of PEP. These two residues are C-terminal to the conventional proline-rich SH3 domain recognition sequence of PEP. This interaction is required in addition to the classic polyproline helix (PPII) recognition by the Csk-SH3 domain for the association between Csk and PEP in vivo. NMR relaxation analysis suggests that Csk-SH3 has different dynamic properties in the various subsites important for peptide recognition.