STAR : Predicting recombination sites from amino acid sequence

Background Designing novel proteins with site-directed recombination has enormous prospects. By locating effective recombination sites for swapping sequence parts, the probability that hybrid sequences have the desired properties is increased dramatically. The prohibitive requirements for applying current tools led us to investigate machine learning to assist in finding useful recombination sites from amino acid sequence alone. Results We present STAR, Site Targeted Amino acid Recombination predictor, which produces a score indicating the structural disruption caused by recombination, for each position in an amino acid sequence. Example predictions contrasted with those of alternative tools, illustrate STAR'S utility to assist in determining useful recombination sites. Overall, the correlation coefficient between the output of the experimentally validated protein design algorithm SCHEMA and the prediction of STAR is very high (0.89). Conclusion STAR allows the user to explore useful recombination sites in amino acid sequences with unknown structure and unknown evolutionary origin. The predictor service is available from http://pprowler.itee.uq.edu.au/star.

Ionized Gas towards Galactic Centre-Constraints from Low-Frequency Recombination Lines

Observations of the H272α recombination line towards the galactic centre show features near VLSR= 0, -50 and + 36 kms-1 . We have combined the parameters of these features with the available -166 measurements to obtain the properties of the ionized gas present along the line of sight and also in the -3 kpc arm-. For the line-of-sight ionized gas we get an electron density around 7 cm-3 and a pathlength through it 10-60 pc. The emission measure and the electron temperature are in the range 500-2900 pc cm-6 and 2000-6000 - respectively. The ionized gas in the 3 kpc arm has an electron density of 30 cm-3 and extends over 9 pc along the line of sight if we assume an electron temperature of 104 K. Using the available upper limit to the intensity of the H351α recombination line, we show that the distributed ionized gas responsible for the dispersion of pulsar signals should have a temperature > 4500 - and a minimum filling factor of 20 per cent. We also show that recombination lines from the -warm ionized- gas proposed by McKee & Ostriker (1977) should be detectable in the frequency range 100-150 MHz towards the galactic centre with the sensitivity available at present.

Genetic analysis of foot-and-mouth disease virus serotype A of Indian origin and detection of positive selection and recombination in leader protease- and capsid-coding regions

The leader protease (L-pro) and capsid-coding sequences (P1) constitute approximately 3 kb of the foot-and-mouth disease virus (FMDV). We studied the phylogenetic relationship of 46 FMDV serotype A isolates of Indian origin collected during the period 1968-2005 and also eight vaccine strains using the neighbour-joining tree and Bayesian tree methods. The viruses were categorized under three major groups - Asian, Euro-South American and European. The Indian isolates formed a distinct genetic group among the Asian isolates. The Indian isolates were further classified into different genetic subgroups (<5% divergence). Post-1995 isolates were divided into two subgroups while a few isolates which originated in the year 2005 from Andhra Pradesh formed a separate group. These isolates were closely related to the isolates of the 1970s. The FMDV isolates seem to undergo reverse mutation or onvergent evolution wherein sequences identical to the ancestors are present in the isolates in circulation. The eight vaccine strains included in the study were not related to each other and belonged to different genetic groups. Recombination was detected in the L-pro region in one isolate (A IND 20/82) and in the VP1 coding 1D region in another isolate (A RAJ 21/96). Positive selection was identified at aa positions 23 in the L-pro (P<0.05; 0.046*) and at aa 171 in the capsid protein VP1 (P<0.01; 0.003**).

Evidence that dicot-infecting mastreviruses are particularly prone to inter-species recombination and have likely been circulating in Australia for longer than in Africa and the Middle East

Viruses of the genus Mastrevirus (family Geminiviridae) are transmitted by leafhoppers and infect either mono- or dicotyledonous plants. Here we have determined the full length sequences of 49 dicot-infecting mastrevirus isolates sampled in Australia, Eritrea, India, Iran, Pakistan, Syria, Turkey and Yemen. Comprehensive analysis of all available dicot-infecting mastrevirus sequences showed the diversity of these viruses in Australia to be greater than in the rest of their known range, consistent with earlier studies, and that, in contrast with the situation in monocot-infecting mastreviruses, detected inter-species recombination events outnumbered intra-species recombination events. Consistent with Australia having the greatest diversity of known dicot-infecting mastreviruses phylogeographic analyses indicating the most plausible scheme for the spread of these viruses to their present locations, suggest that most recent common ancestor of these viruses is likely nearer Australia than it is to the other regions investigated.

Molecular diversity of Chickpea chlorotic dwarf virus in Sudan: High rates of intra-species recombination – a driving force in the emergence of new strains

In Sudan Chickpea chlorotic dwarf virus (CpCDV, genus Mastrevirus, family Geminiviridae) is an important pathogen of pulses that are grown both for local consumption, and for export. Although a few studies have characterised CpCDV genomes from countries in the Middle East, Africa and the Indian subcontinent, little is known about CpCDV diversity in any of the major chickpea production areas in these regions. Here we analyse the diversity of 146 CpCDV isolates characterised from pulses collected across the chickpea growing regions of Sudan. Although we find that seven of the twelve known CpCDV strains are present within the country, strain CpCDV-H alone accounted for ∼73% of the infections analysed. Additionally we identified four new strains (CpCDV-M, -N, -O and -P) and show that recombination has played a significant role in the diversification of CpCDV, at least in this region. Accounting for observed recombination events, we use the large amounts of data generated here to compare patterns of natural selection within protein coding regions of CpCDV and other dicot-infecting mastrevirus species.

Structure of Mycobacterium tuberculosis RuvA, a protein involved in recombination

The process of recombinational repair is crucial for maintaining genomic integrity and generating biological diversity. In association with RuvB and RuvC, RuvA plays a central role in processing and resolving Holliday junctions, which are a critical intermediate in homologous recombination. Here, the cloning, purification and structure determination of the RuvA protein from Mycobacterium tuberculosis (MtRuvA) are reported. Analysis of the structure and comparison with other known RuvA proteins reveal an octameric state with conserved subunit-subunit interaction surfaces, indicating the requirement of octamer formation for biological activity. A detailed analysis of plasticity in the RuvA molecules has led to insights into the invariant and variable regions, thus providing a framework for understanding regional flexibility in various aspects of RuvA function.

Phage lambda beta protein, a component of general recombination, is associated with host ribosomal S1 protein

We have purified phage lambda beta protein produced by a recombinant plasmid carrying bet gene and confirm that it forms a complex with a protein of relative molecular mass 70 kDa. Therefore, beta protein, a component of general genetic recombination, is associated with two functionally diverse complexes; one containing exonuclease and the other 70 kDa protein. Using a number of independent methods, we show that 70 kDa protein is the ribosomal S1 protein of E. coli. Further, the association of 70 kDa protein with beta protein is biologically significant, as the former inhibits joining of the terminal ends of lambda chromosome and renaturation of complementary single stranded DNA promoted by the latter. More importantly, these findings initiate an understanding of an important mode of host- virus interaction in general with specific implication(s) in homologous genetic recombination.

The homologous recombination system of phage lambda. Pairing activities of beta protein

The red genes of phage lambda specify two proteins, exonuclease and beta protein, which are essential for its general genetic recombination in recA- cells. These proteins seem to occur in vivo as an equimolar complex. In addition, beta protein forms a complex with another polypeptide, probably of phage origin, of Mr 70,000. The 70-kDa protein appears to be neither a precursor nor an aggregated form of either exonuclease or beta protein, since antibodies directed against the latter two proteins failed to react with 70-kDa protein on Ouchterlony double diffusion analysis. beta protein promotes Mg2+-dependent renaturation of complementary strands (Kmiec, E., and Holloman, W. K. (1981) J. Biol. Chem. 256, 12636-12639). To look for other pairing activities of beta protein, we developed methods of purification to free it of associated exonuclease. Exonuclease-free beta protein appeared unable to cause the pairing of a single strand with duplex DNA; however, like Escherichia coli single strand binding protein (SSB), beta protein stimulated formation of joint molecules by recA protein from linear duplex DNA and homologous circular single strands. Like recA protein, but unlike SSB, beta protein promoted the joining of the complementary single-stranded ends of phage lambda DNA. beta protein specifically protected single-stranded DNA from digestion by pancreatic DNase. The half-time for renaturation catalyzed by beta protein was independent of DNA concentration, unlike renaturation promoted by SSB and spontaneous renaturation, which are second order reactions. Thus, beta protein resembles recA protein in its ability to bring single-stranded DNA molecules together and resembles SSB in its ability to reduce secondary structure in single-stranded DNA.

Recombination and trapping in n-type cobalt-doped germanium

The role of cobalt centers in promoting the recombination and trapping processes in n-type germanium has been investigated. Data on lifetime measurements carried out by the steadystate photoconductivity and photo-magneto-electric methods in the temperature range 145 to 300°K on n-type germanium samples containing cobalt in the concentration range 1·1013 to 5.·014/cm3 are presented. The results are analysed on the basis of Sah-Shockley's multi-level formula to yield the capture cross-sections Sp= (hole capture cross-section at doubly negatively charged center) and Sn-(electron capture cross-section at singly negatively charged center) and temperature dependence. It is found that Sp= is (22 ± 6). 10-16 cm2 and Sn- is ∼ 0·1. 10-16 cm2 at 145°K. Sp= varies (n = 3·5 to 4·5) in the range 145-220°K; above 225°K the index 'n' tends to a smaller value. Sn- is practically temperature independent below 180°K and increases with increase of temperature above 180°K. The value of Sp= and its temperature variation lead one to the conclusion that during capture at attractive centers, the phonon cascade mechanism is responsible for the dissipation of the recombination energy.

Electron-hole recombination in cobalt-doped p-type germanium

The recombination properties of cobalt centers in p-type germanium containing cobalt in the concentration range 1014 to 1016 atoms/cm3 have been investigated. The measurement of lifetime has been carried out by steady-state photoconductivity and photo-magneto-electric methods in the temperature range 145 to 300°K. The cross-sections Sno (electron capture cross-section at neutral centers). Sn- (electron capture cross-section at singly negatively charged centers) and their temperature variations have been estimated by the analysis of the lifetime data on the basis of Sah-Shockley's multi-level formula. The value of Sno is (15±5).10-16 cm2 and is temperature independent. The value of Sn- is ≈4·10-16 cm2 around 225°K and it increases with increase of temperature. The possible mechanisms for capture at neutral and repulsive centers are discussed and a summary of the capture cross-sections for cobalt centers is given. A comparison of the cross-section values of cobalt and their temperature variations with those of the related impurities-manganese, iron and nickel-in germanium has been made.

Cytosines, but Not Purines, Determine Recombination Activating Gene (RAG)-induced Breaks on Heteroduplex DNA Structures

The sequence specificity of the recombination activating gene (RAG) complex during V(D)J recombination has been well studied. RAGs can also act as structure-specific nuclease; however, little is known about the mechanism of its action. Here, we show that in addition to DNA structure, sequence dictates the pattern and efficiency of RAG cleavage on altered DNA structures. Cytosine nucleotides are preferentially nicked by RAGs when present at single-stranded regions of heteroduplex DNA. Although unpaired thymine nucleotides are also nicked, the efficiency is many fold weaker. Induction of single- or double-strand breaks by RAGs depends on the position of cytosines and whether it is present on one or both of the strands. Interestingly, RAGs are unable to induce breaks when adenine or guanine nucleotides are present at single-strand regions. The nucleotide present immediately next to the bubble sequence could also affect RAG cleavage. Hence, we propose “C(d)C(S)C(S)” (d, double-stranded; s, single-stranded) as a consensus sequence for RAG-induced breaks at single-/double-strand DNA transitions. Such a consensus sequence motif is useful for explaining RAG cleavage on other types of DNA structures described in the literature. Therefore, the mechanism of RAG cleavage described here could explain facets of chromosomal rearrangements specific to lymphoid tissues leading to genomic instability.

Starburst in the ultraluminous galaxy Arp 220: Constraints from observations of radio recombination lines and continuum

We present observations of radio recombination lines (RRL) from the starburst galaxy Arp 220 at 8.1 GHz (H92 alpha) and 1.4 GHz (H167 alpha and H165 alpha) and at 84 GHz (H42 alpha), 96 GHz (H40 alpha) and 207 GHz (H31 alpha) using the Very Large Array and the IRAM 30 m telescope, respectively. RRLs were detected at all the frequencies except 1.4 GHz, where a sensitive upper limit was obtained. We also present continuum flux measurements at these frequencies as well as at 327 MHz made with the VLA. The continuum spectrum, which has a spectral index alpha similar to -0.6 (S-nu proportional to nu(alpha)) between 5 and 10 GHz, shows a break near 1.5 GHz, a prominent turnover below 500 MHz, and a flatter spectral index above 50 GHz. We show that a model with three components of ionized gas with different densities and area covering factors can consistently explain both RRL and continuum data. The total mass of ionized gas in the three components is 3.2 x 10(7) M., requiring 3 x 10(5) O5 stars with a total Lyman continuum production rate N-Lyc similar to 1.3 x 10(55) photons s(-1). The ratio of the expected to observed Br alpha and Br gamma fluxes implies a dust extinction A(V) similar to 45 mag. The derived Lyman continuum photon production rate implies a continuous star formation rate (SFR) averaged over the lifetime of OB stars of similar to 240 M yr(-1). The Lyman continuum photon Production rate of similar to 3% associated with the high-density H II regions implies a similar SFR at recent epochs (t < 10(5) yr). An alternative model of high-density gas, which cannot be excluded on the basis of the available data, predicts 10 times higher SFR at recent epochs. If confirmed, this model implies that star formation in Arp 220 consists of multiple starbursts of very high SFR (few times 10(3) M. yr(-1)) and short duration (similar to 10(5) yr). The similarity of IR excess, L-IR/L-Ly alpha similar to 24, in Arp 220 to values observed in starburst galaxies shows that most of the high luminosity of Arp 220 is due to the ongoing starburst rather than to a hidden active galactic nucleus (AGN). A comparison of the IR excesses in Arp 220, the Galaxy, and M33 indicates that the starburst in Arp 220 has an initial mass function that is similar to that in normal galaxies and has a duration longer than 107 yr. If there was no infall of gas during this period, then the star formation efficiency (SFE) in Arp 220 is similar to 50%. The high SFR and SFE in Arp 220 is consistent with their known dependences on mass and density of gas in star-forming regions of normal galaxies.

Surface recombination at semiconductor electrodes: a single level study

The recombination and the faradaic fluxes are shown to be sensitive to the location of a single level recombination center, when it is located near the band edges. As the surface level is shifted deeper into the band gap from either of the band edges, the back emission terms are dominated by electron capture and hole capture terms, and the occupancy of the surface level is no longer determined by its location in the band gap. However, when one of the back emission terms determines the surface state occupancy, it is shown that there exists a simple relation between the value of the surface level and the recombination and the faradaic fluxes respectively. Expressions to this effect are derived and verified in the case of the recombination flux, which characterized by the potential at which it attains its maximum value. For the faradaic flux the results are qualitative.