The binding of pentaammineruthenium (III) to RNase B and RNase A+d(pA)4 in the crystalline state

The binding of pentaammineruthenium (III) to ribonuclease A and B both free and complexed with d(pA)₄ has been examined in the crystalline state through the application of X-ray diffraction and difference Fourier techniques. In crystals of native RNase B, the reagent was observed to have many binding sites, some entirely electrostatic in nature and others consistent with coordination to histidine residues. The primary histidine in the latter case was 105 with 119 also partially substituted. In crystals of RNase A+d(pA)₄ complex only a single, extremely strong site of substitution was observed, and this was 2.4 Å from the native position of the imidazole ring of histidine 105. Thus, the results of these X-ray diffraction studies appear to be quite consistent with the findings of earlier NMR studies and with the results obtained in crystals of the gene 5 DNA binding protein.     

The N-Terminal Region of the RecU Holliday Junction Resolvase Is Essential for Homologous Recombination

The RecU Holliday junction (HJ)-resolving enzyme is highly conserved in the Firmicutes phylum of bacteria. In Bacillus subtilis, the recU gene has two putative initiation codons, at positions 1 and 33. In rec⁺ cells, only the full-length RecU polypeptide (206 residues, 23.9 kDa) was detected even after different stress treatments. To address the relevance of the flexible N-terminus, we constructed mutant variants. Experiments in vivo revealed that recUΔ1-32 (which initiates at Met33 and encodes RecUΔ1-32) and recU31 (the conserved Arg31 residue was substituted with alanine to give RecUR31A) are genuine RecU mutants, rendering cells impaired in DNA repair and chromosomal segregation. RecU has three activities: It (i) cleaves HJs, (ii) anneals complementary strands and (iii) modulates RecA activities. RecUR31A binds and cleaves HJ DNA in vitro as efficiently as wild-type RecU, but RuvB·ATPγS·Mg²⁺ fails to stimulate the RecUR31A cleavage reaction. In contrast, RecUΔ1-32 forms unstable complexes with DNA and fails to cleave HJs. RecU and its variants are capable of promoting DNA strand annealing and exert a negative effect on deoxy-ATP-dependent RecA-mediated DNA strand exchange. This study shows that the flexible N-terminus of RecU is essential for protein activity.     

Roles of non‐native hydrogen‐bonding interaction in helix‐coil transition of a single polypeptide as revealed by comparison between Gō‐like and non‐Gō models

To explore the role of non‐native interactions in the helix‐coil transition, a detailed comparison between a Gō‐like model and a non‐Gō model has been performed via lattice Monte Carlo simulations. Only native hydrogen bonding interactions occur in the Gō‐like model, and the non‐native ones with sequence interval more than 4 is also included into the non‐Gō model. Some significant differences between the results from those two models have been found. The non‐native hydrogen bonds were found most populated at temperature around the helix‐coil transition. The rearrangement of non‐native hydrogen bonds into native ones in the formation of α‐helix leads to the increase of susceptibility of chain conformation, and even two peaks of susceptibility of radius of gyration versus temperature exist in the case of non‐Gō model for a non‐short peptide, while just a single peak exists in the case of Gō model for a single polypeptide chain with various chain lengths. The non‐native hydrogen bonds have complicated the temperature‐dependence of Zimm‐Bragg nucleation constant. The increase of relative probability of non‐native hydrogen bonding for long polypeptide chains leads to non‐monotonous chain length effect on the transition temperature. Proteins 2010. © 2010 Wiley‐Liss, Inc.     

Der Einfluß von Rapsextraktionsschrot und Methylthiourazil auf die Schilddrüsensekretionsrate von Hühnern nach kurzfristiger Fütterungsdauer

Three experiments were carried out with male broiler chickens reared from day‐ old to 6 weeks of age on semi‐purified diets containing 10% fresh (Expt. 1 and 3) or oxidized (Expt. 2) re‐esterified triglycerides with a fatty acid composition similar to that of soya bean oil containing increasing concentrations of either a mixture of d‐α‐, γ‐, δ‐tocopherylacetate (d‐tocopherols) of natural source or dl‐α‐ tocopheryl acetate (dl‐tocopherol). In Expt. 1 and 2 the mixture of d‐tocopherols consisted of 35.7% d‐α‐, 45.3% d‐γ‐ and 19.0% d‐δ‐, while in Expt. 3 the distribution was 25.3% d‐α‐, 28.1% d‐γ‐and 10.8% d‐γ‐ in 35.8% re‐ esterified triglycerides. The relative biopotency of d‐α‐: γ‐: δ‐tocopherol was anticipated to be 100: 25: 1, whereas that of dl‐a‐tocopherol was 74% relative to d‐ a‐tocopherol. The experiments demonstrate that the results obtained for the biological activity depend on the response parameters chosen. With respect to gain in weight, feed conversion, relative organ weight, packed cell volume (PCV), ELP (erythrocytelipidperoxidation), plasma activities of glutamate‐oxaloacetate‐transaminase (GOT), creatine kinase (CK) and glutathione peroxidase (GSH‐Px) and plasma Na⁺ concentration, the mixture of natural source tocopherols was identical to that of dl‐α‐tocopheryl acetate, although the concentration of a‐ tocopherol was only about one third of that of dl‐α‐tocopherol. Differences between natural source and synthetic tocopherols were expectedly observed with respect to plasma concentrations of α‐, γ‐, δ‐tocopherol. Differences between the two forms as to muscular dystrophy, in vitro haemolysis and potassium concentration in plasma were ambigous. It is suggested that the function of d‐α‐, γ‐, δ‐tocopherol in erythrocyte fragility and skeletal muscle structure should be compared to that of dl‐α‐tocopherol in future investigations.     

Synthesis of New Homo and Heterodimers of 2′,3′-Dideoxyinosine (ddi) Using Ester Linkages

Abstract

A series of new homo and heterodimers of ddI has been synthesized. A glutarate diester spacer was used to covalently couple ddI onto ddI, AZT or d4T.     

Enhanced operation of femtosecond lasers and applications in cell transfection

In this work we present a review and discussion on the enhancement of femtosecond (fs) lasers for use within biophotonics with a particular focus on their use in optical transfection techniques. We describe the broad range of source options now available for the generation of femtosecond pulses before briefly reviewing the application of fs laser in optical transfection studies. We show that major performance enhancements may be obtained by optimising the spatial and temporal performance of the laser source before considering possible future directions in this field. In relation to optical transfection we describe how such laser sources initiate a multiphoton process to permeate the cell membrane in a transient fashion. We look at aspects of this technique including the ability to combine transfection with optical trapping. For future implementation of such transfection we explore the role of new sources and “nondiffracting” light fields. (© 2008 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)