A generalized conformational energy function of DNA derived from molecular dynamics simulations

Proteins recognize DNA sequences by two different mechanisms. The first is direct readout, in which recognition is mediated by direct interactions between the protein and the DNA bases. The second is indirect readout, which is caused by the dependence of conformation and the deformability of the DNA structure on the sequence. Various energy functions have been proposed to evaluate the contribution of indirect readout to the free-energy changes in complex formations. We developed a new generalized energy function to estimate the dependence of the deformability of DNA on the sequence. This function was derived from molecular dynamics simulations previously conducted on B-DNA dodecamers, each of which had one possible tetramer sequence embedded at its center. By taking the logarithm of the probability distribution function (PDF) for the base-step parameters of the central base-pair step of the tetramer, its ability to distinguish the native sequence from random ones was superior to that with the previous method that approximated the energy function in harmonic form. From a comparison of the energy profiles calculated with these two methods, we found that the harmonic approximation caused significant errors in the conformational energies of the tetramers that adopted multiple stable conformations.     

Characterization of monocyte differentiation-inducing (MDI) factors derived from human fetal membrane chorion cells undergoing apoptosis after influenza virus infection

Influenza virus infection during pregnancy has been implicated as one of cause of premature delivery, abortion and stillbirth. We have reported that cultured human fetal membrane chorion cells undergoing apoptosis by influenza virus infection secrete unidentified heat-stable monocyte differentiation-inducing (MDI) factors. In this study, cellular, biological and immunochemical characteristics of MDI factors were investigated using human monocytic leukemia THP-1 cells by nitroblue tetrazolium reduction and cell adhesion assays. The treatment of THP-1 cells with culture supernatants from the influenza virus-infected chorion cells induced the nitroblue tetrazolium reduction ability, which was inhibited by the addition of superoxide dismutase and diphenyleneiodonium chloride, an inhibitor for reduced nicotinamide adenine dinucleotide phosphate oxidase. The phenomenon was also observed in human peripheral blood monocytes and histiocytic leukemia U937 cells, but not in promyelocytic leukemia HL-60 cells. The induction of nitroblue tetrazolium reduction and adhesion abilities in THP-1 cells was closely correlated with the concentrations of interleukin-6 protein in the culture supernatants. These abilities were inhibited to approximately 60% by the addition of antibodies against interleukin-6, or alpha-chain (gp80) or beta-chain (gp130) of IL-6 receptor. The induction of nitroblue tetrazolium reduction was increased by the addition of supernatants from amniochorion tissue cultures after influenza virus infection. These results indicate that chorion cell-derived interleukin-6 is partly responsible for monocyte differentiation to macrophages capable of generating superoxide anion. It is possible that these pathways represent part of the mechanism for birth complications associated with intrauterine influenza infection in pregnancy.     

Contribution of a single hydrogen bond between betaHis81 of MHC class II I-E(k) and the bound peptide to the pH-dependent thermal stability

To determine the energetic contribution of the hydrogen bond between betaHis81 of the major histocompatibility complex class II (MHC II) molecule, I-E(k), and the bound hemoglobin peptide (Hb), we analyzed the thermal stability of the hydrogen bond-disrupted mutant, I-E(k)-Hb betaH81Y, in which the betaHis81 residue was replaced with Tyr, by differential scanning calorimetry. The thermal stability of the I-E(k)-Hb betaH81Y mutant was lower than that of the I-E(k)-Hb wild-type, mainly due to the decreased enthalpy change. The difference in the denaturation temperature of the I-E(k)-Hb betaH81Y mutant as compared with that of the I-E(k)-Hb wild-type at pH 5.5 was only slightly smaller than that at pH 7.4, in agreement with the increased stability at an acidic pH, a unique characteristic of MHC II. Thus, the hydrogen bond contributed by betaHis81 is critical for peptide binding, and is independent of pH, which can alter the hydrophilicity of the His residue.     

Interrelations between the efficiency of translation start sites and other sequence features of yeast mRNAs

The translation start site (TSS) plays an important role in the control of the translational efficiency and cytoplasmic stability of eukaryotic mRNAs. The efficiency of TSS recognition is known to be influenced by sequence context, and mRNAs with "weak" TSSs are relatively abundant. We analyzed a sample of 4113 yeast genes in a search for features that might serve to compensate for the inefficient recognition of "weak" TSSs by initiating ribosomes. The first feature found to correlate with variations in TSS strength is differences in the stability of secondary structure upstream and downstream of the start AUG codon. The second feature concerns the characteristics of AUG triplets found at the beginning of the coding sequence, i.e., downstream of the predicted TSS. In particular, the proximal downstream AUG lies in frame with the CDS significantly more often if the TSS itself is located in a "weak" context. The accuracy of TSS annotation, the possibility of polypeptide heterogeneity due to the use of alternative downstream AUGs, and the influence of related features of mRNA sequences are discussed.     

Importance of mutant position in Ramachandran plot for predicting protein stability of surface mutations

Understanding the mechanisms by which mutations affect protein stability is one of the most important problems in molecular biology. In this work, we analyzed the relationship between changes in protein stability caused by surface mutations and changes in 49 physicochemical, energetic, and conformational properties of amino acid residues. We found that the hydration entropy was the major contributor to the stability of surface mutations in helical segments; other properties responsible for size and volume of molecule also correlated significantly with stability. Classification of coil mutations based on their locations in the (phi-psi) map improved the correlation significantly, demonstrating the existence of a relationship between stability and strain energy, which indicates that the role of strain energy is very important for the stability of surface mutations. We observed that the inclusion of sequence and structural information raised the correlation, indicating the influence of surrounding residues on the stability of surface mutations. Further, we examined the previously reported "inverse relationship" between stability and hydrophobicity, and observed that the inverse hydrophobic effect was generally applicable only to coil mutations. The present study leads to a simple method for predicting protein stability changes caused by amino acid substitutions, which will be useful for protein engineering in designing novel proteins with increased stability and altered function.     

A backflow of electrons around photosystem II in Chlorella cells.

A study was made with a modulated oxygen electrode of the effect of variations of oxygen concentration on photosynthetic oxygen evolution from algal cells. When Chlorella vulgaris is examined with a modulated 650 nm light at 22 degrees C, both the oxygen yield and the phase lag between the modulated oxygen signal and the light modulations have virtually constant values between 800 and 120 ergs . cm-1 . s-1 if the bathing medium is in equilibrium with the air. Similar results are obtained at 32 degrees C between 1600 and 120 ergs . cm-2 . s-1. Under anaerobic conditions both the oxygen yield and the phase lag decrease if the light intensity is lowered below about 500 ergs . cm-2 . s-1 at 22 degrees C or about 1000 ergs . cm-2 . s-1 at 32 degrees C. A modulated 706 nm beam also gives rise to these phenomena but only at significantly lower rates of oxygen evolution. The cells of Anacystis nidulans and Porphyridium cruentum appear to react in the same way to anaerobic conditions as C. vulgaris. An examination of possible mechanisms to explain these results was performed using a computer simulation of photosynthetic electron transport. The simulation suggests that a backflow of electrons from a redox pool between the Photosystems to the rate-limiting reaction between Photosystem II and the water-splitting act can cause a decrease in oxygen yield and phase lag. If the pool between the Photosystems is in a very reduced state a significant cyclic flow is expected, whereas if the pool is largely oxidized little or no cyclic flow should occur. It is shown that the effects of 706 nm illumination and removal of oxygen can be interpreted in accordance with these proposals. Since a partial inhibition of oxygen evolution by 3-(3.4-dichlorophenyl)-1,1-dimethylurea (10(-8) M) magnifies the decreases in oxygen yield and phase lag, it is proposed that the pool which cycles back electrons is in front of the site of 3-(3,4-dichlorophenyl)-1,1-dimethylurea inhibition and is probably the initial electron acceptor pool after Photosystem II.     

Efficiency of electron transport processes

Efficiency of electron transport along the linear chain of molecules was investigated from a dynamic viewpoint. It was proposed that two kinds of efficiency are important for electron transport; one is energy efficiency, the other quantum efficiency. In this paper, these two efficiencies are defined for a linear chain system and the correlation between these quantities and the arrangement of various electron transfers is investigated. The optimization of energy and quantum efficiency is found to set different conditions on the arrangement of the rate constants of electron transfer, and there is strong correlation between neighboring electron transfers. In order to maximize both efficiencies, the rate constants of forward and backward transfers of electrons should be bounded by one another in a limited range. In particular, when there are some bypass reactions on the linear chain, as is the case for photosynthesis and respiration, the rate of the backward transfer should be the same order of magnitude as that of the next forward transfer. The present results are applied to some biological processes. In the early stage of photosynthetic electron transfer it seems that quantum efficiency is more important than energy efficiency. The quantum efficiency is close to unity, whereas a considerable part of the free energy is wasted as heat during the primary electron transfers. On the other hand, in the slower electron transfer processes in photosynthesis and respiration, which take place mostly near equilibrium, the energy efficiency seems to be more important than the quantum efficiency. The relation of these properties to biological function is discussed.     

The role of alternative translation start sites in the generation of human protein diversity

According to the scanning model, 40S ribosomal subunits initiate translation at the first (5 proximal) AUG codon they encounter. However, if the first AUG is in a suboptimal context, it may not be recognized, and translation can then initiate at downstream AUG(s). In this way, a single RNA can produce several variant products. Earlier experiments suggested that some of these additional protein variants might be functionally important. We have analysed human mRNAs that have AUG triplets in 5 untranslated regions and mRNAs in which the annotated translational start codon is located in a suboptimal context. It was found that 3% of human mRNAs have the potential to encode N-terminally extended variants of the annotated proteins and 12% could code for N-truncated variants. The predicted subcellular localizations of these protein variants were compared: 31% of the N-extended proteins and 30% of the N-truncated proteins were predicted to localize to subcellular compartments that differed from those targeted by the annotated protein forms. These results suggest that additional AUGs may frequently be exploited for the synthesis of proteins that possess novel functional properties.Electronic Supplementary Material Supplementary material is available for this article at     

PSSM-based prediction of DNA binding sites in proteins

Background  

Detection of DNA-binding sites in proteins is of enormous interest for technologies targeting gene regulation and manipulation. We have previously shown that a residue and its sequence neighbor information can be used to predict DNA-binding candidates in a protein sequence. This sequence-based prediction method is applicable even if no sequence homology with a previously known DNA-binding protein is observed. Here we implement a neural network based algorithm to utilize evolutionary information of amino acid sequences in terms of their position specific scoring matrices (PSSMs) for a better prediction of DNA-binding sites.