Network of long-range concerted chemical shift displacements upon ligand binding to human angiogenin

Molecular recognition models of both induced fit and conformational selection rely on coupled networks of flexible residues and/or structural rearrangements to promote protein function. While the atomic details of these motional events still remain elusive, members of the pancreatic ribonuclease superfamily were previously shown to depend on subtle conformational heterogeneity for optimal catalytic function. Human angiogenin, a structural homologue of bovine pancreatic RNase A, induces blood vessel formation and relies on a weak yet functionally mandatory ribonucleolytic activity to promote neovascularization. Here, we use the NMR chemical shift projection analysis (CHESPA) to clarify the mechanism of ligand binding in human angiogenin, further providing information on long-range intramolecular residue networks potentially involved in the function of this enzyme. We identify two main clusters of residue networks displaying correlated linear chemical shift trajectories upon binding of substrate fragments to the purine- and pyrimidine-specific subsites of the catalytic cleft. A large correlated residue network clusters in the region corresponding to the V₁ domain, a site generally associated with the angiogenic response and structural stability of the enzyme. Another correlated network (residues 40–42) negatively affects the catalytic activity but also increases the angiogenic activity. ¹⁵N-CPMG relaxation dispersion experiments could not reveal the existence of millisecond timescale conformational exchange in this enzyme, a lack of flexibility supported by the very low-binding affinities and catalytic activity of angiogenin. Altogether, the current report potentially highlights the existence of long-range dynamic reorganization of the structure upon distinct subsite binding events in human angiogenin.     

The influence of sunlight on the localized corrosion of UNS S31600 in natural seawater

Tests were conducted on the performance of UNS S31600 stainless steel (SS) in a natural day/night cycle vs full darkness under conditions of natural marine biofilm accumulation. In quiescent flowing seawater tests in the laboratory as well as under natural immersion in the sea, diffuse sunlight (～10% of natural) counteracted the influence of marine biofilms and produced substantial inhibition of the corrosion of SS. Thus, the probabilities (percentage attack) and propagation rates (depths of attack) in multiple crevice tests were substantially lower in the day/night cycle than in the dark. A benefit was also observed for welded SS in terms of the time to corrosion initiation and the mass loss. SS in the passive state showed broader passive regions, well-defined breakdown potentials and markedly smaller anodic and cathodic current densities under the diurnal cycle. The overall reduction in corrosion is attributed to a combination of electrochemical photoinhibition and simultaneous photoinactivation of microbially mediated metal redox reactions linked to cathodic kinetics. These data offer fresh insights into the behaviour of SS under practical seawater situations and the proposed potential use of illumination in the mitigation of biologically influenced consequences.     

Proteome Composition in Plasmodium falciparum: Higher Usage of GC-Rich Nonsynonymous Codons in Highly Expressed Genes

The parasite Plasmodium falciparum, responsible for the most deadly form of human malaria, is one of the extremely AT-rich genomes sequenced so far and known to possess many atypical characteristics. Using multivariate statistical approaches, the present study analyzes the amino acid usage pattern in 5038 annotated protein-coding sequences in P. falciparum clone 3D7. The amino acid composition of individual proteins, though dominated by the directional mutational pressure, exhibits wide variation across the proteome. The Asn content, expression level, mean molecular weight, hydropathy, and aromaticity are found to be the major sources of variation in amino acid usage. At all stages of development, frequencies of residues encoded by GC-rich codons such as Gly, Ala, Arg, and Pro increase significantly in the products of the highly expressed genes. Investigation of nucleotide substitution patterns in P. falciparum and other Plasmodium species reveals that the nonsynonymous sites of highly expressed genes are more conserved than those of the lowly expressed ones, though for synonymous sites, the reverse is true. The highly expressed genes are, therefore, expected to be closer to their putative ancestral state in amino acid composition, and a plausible reason for their sequences being GC-rich at nonsynonymous codon positions could be that their ancestral state was less AT-biased. Negative correlation of the expression level of proteins with respective molecular weights supports the notion that P. falciparum, in spite of its intracellular parasitic lifestyle, follows the principle of cost minimization. [Reviewing Editor : Dr. Richard Kliman]     

Thermo-optically Induced Reorganizations in the Main Light Harvesting Antenna of Plants. I. Non-Arrhenius Type of Temperature Dependence and Linear Light-intensity Dependencies

Thermo-optically induced structural reorganizations have earlier been identified in isolated LHCII, the main chlorophyll a/b light harvesting complexes of Photosystem II, and in granal thylakoid membranes [Cseh et al. (2000) Biochemistry 39: 15250–15257; Garab et al. (2002) Biochemistry 41: 15121–15129]. According to the thermo-optic mechanism, structural changes can be induced by fast, local thermal transients due to the dissipation of excess excitation energy. In this paper, we analyze the temperature and light-intensity dependencies of thermo-optically induced reversible and irreversible reorganizations in the chiral macrodomains of lamellar aggregates of isolated LHCII and of granal thylakoid membranes. We show that these structural changes exhibit non-Arrhenius type of temperature dependencies, which originate from the ‘combination’ of the ambient temperature and the local thermal transient. The experimental data can satisfactorily be simulated with the aid of a simple mathematical model based on the thermo-optic effect. The model also predicts, in good accordance with experimental data published earlier and presented in this paper, that the reorganizations depend linearly on the intensity of the excess light, a unique property that is probably important in light adaptation and photoprotection of plants.     

Features of ribosome-peptidyl-tRNA interactions essential for tryptophan induction of tna operon expression

Certain nascent peptide sequences, when within the ribosomal exit tunnel, can inhibit translation termination and/or peptide elongation. The 24 residue leader peptidyl-tRNA of the tna operon of E. coli, TnaC-tRNA(Pro), in the presence of excess tryptophan, resists cleavage at the tnaC stop codon. TnaC residue Trp12 is crucial for this inhibition. The approximate location of Trp12 in the exit tunnel was determined by crosslinking Lys11 of TnaC-tRNA(Pro) to nucleotide A750 of 23S rRNA. Methylation of nucleotide A788 of 23S rRNA was reduced by the presence of Trp12 in TnaC-tRNA(Pro), implying A788 displacement. Inserting an adenylate at position 751, or introducing the change U2609C in 23S rRNA or the change K90H or K90W in ribosomal protein L22, virtually eliminated tryptophan induction. These modified and mutated regions are mostly located near the putative site occupied by Trp12 of TnaC-tRNA(Pro). These findings identify features of the ribosomal exit tunnel essential for tna operon induction.     

Roles of the threonine 407, aspartic acid 417, and threonine 419 residues in P450 2B1 in metabolism

We have previously observed that the quadruple (S407T-N417D-A419T-K473M) and triple (S407T-N17D-A419T) mutants of the chimeric construct of P450 2B1/2B2 do not undergo mechanism-based inactivation by 17alpha-ethynylestradiol (17EE) and tert-butyl 1-methyl-2-propynyl ether (tBMP). The ability of these mutants to metabolize 17EE, benzphetamine, and testosterone has been investigated. The profile for 17EE metabolism by both mutants was characteristic of both wild-types. The two mutants metabolized testosterone to form androstenedione with no formation of the hydroxy products as was seen with both the wild-types. Benzphetamine metabolism by the mutants showed that both mutants exhibited an increased tendency to catalyze demethylation rather than debenzylation. In the presence of the alternate oxidants cumene hydroperoxide and tert-butyl hydroperoxide, the wild-type 2B1 was not inactivated by 17EE. Metabolism of 17EE by 2B1 supported by these alternate oxidants revealed differences in the metabolites that may be related to the inability of 2B1 to be inactivated under these conditions.     

Iopamidol myelography-induced seizures

Iopamidol, a water-soluble contrast agent, has been rarely associated with seizures. We describe a case of generalized tonic-clonic seizure after cervical myelography with iopamidol in a previously healthy young man. In patients presenting with seizures, a history of recent myelography should be considered as an etiology. Iopamidol myelography may be associated with a risk of seizures. Clinicians need to be aware of this complication and inform their patients about such risk.     

Biophysical approaches for studying the integrity and function of tight junctions

Cell-cell adhesion is an extremely important phenomenon as it influences several biologically important processes such as inflammation, cell migration, proliferation, differentiation and even cancer metastasis. Furthermore, proteins involved in cell-cell adhesion are also important from the perspective of facilitating better drug delivery across epithelia. The adhesion forces imparted by proteins involved in cell-cell adhesion have been the focus of research for sometime. However, with the advent of nanotechnological techniques such as the atomic force microscopy (AFM), we can now quantitatively probe these adhesion forces not only at the cellular but also molecular level. Here, we review the structure and function of tight junction proteins, highlighting some mechanistic studies performed to quantify the adhesion occurring between these proteins and where possible their association with human diseases. In particular, we will highlight two important experimental techniques, namely the micropipette step pressure technique and the AFM that allow us to quantify these adhesion forces at both the cellular and molecular levels, respectively.