The 43 residue DNA binding domain of gamma delta resolvase binds adjacent major and minor grooves of DNA.

The carboxyl-terminal domain of gamma delta resolvase binds to each half of the three resolvase binding sites that constitute the recombination site, res. Ethylation inhibition experiments show that the phosphate contacts made by the C-terminal DNA binding domain are similar to those made by intact resolvase, with the exception of a single phosphate at the inside end of each contact region which is contacted solely by the intact resolvase. The DNA binding domain makes essentially identical contacts to all 6 half sites, whereas the intact resolvase makes slightly different contacts to each binding site. Despite its small size, only 43 amino acid residues, the resolvase C-terminal domain interacts with an unusually large segment of DNA. Phosphate contacts extend across an adjacent major and minor groove of DNA and about one third of the circumference around the helix. The minimal binding segment, determined experimentally, is a 12 bp sequence that includes the 9 base pair inverted repeat (common to all half sites), the adjacent 3 base pairs (towards the center of the intact resolvase binding site), and phosphates at both ends.     

Dark Matter of the Biosphere: the Amazing World of Bacteriophage Diversity

Bacteriophages are the most abundant biological entities in the biosphere, and this dynamic and old population is, not surprisingly, highly diverse genetically. Relative to bacterial genomics, phage genomics has advanced slowly, and a higher-resolution picture of the phagosphere is only just emerging. This view reveals substantial diversity even among phages known to infect a common host strain, but the relationships are complex, with mosaic genomic architectures generated by illegitimate recombination over a long period of evolutionary history.     

Imbroglios of viral taxonomy: genetic exchange and failings of phenetic approaches

The practice of classifying organisms into hierarchical groups originated with Aristotle and was codified into nearly immutable biological law by Linnaeus. The heart of taxonomy is the biological species, which forms the foundation for higher levels of classification. Whereas species have long been established among sexual eukaryotes, achieving a meaningful species concept for prokaryotes has been an onerous task and has proven exceedingly difficult for describing viruses and bacteriophages. Moreover, the assembly of viral "species" into higher-order taxonomic groupings has been even more tenuous, since these groupings were based initially on limited numbers of morphological features and more recently on overall genomic similarities. The wealth of nucleotide sequence information that catalyzed a revolution in the taxonomy of free-living organisms necessitates a reevaluation of the concept of viral species, genera, families, and higher levels of classification. Just as microbiologists discarded dubious morphological traits in favor of more accurate molecular yardsticks of evolutionary change, virologists can gain new insight into viral evolution through the rigorous analyses afforded by the molecular phylogenetics of viral genes. For bacteriophages, such dissections of genomic sequences reveal fundamental flaws in the Linnaean paradigm that necessitate a new view of viral evolution, classification, and taxonomy.     

Integration and excision of the Mycobacterium tuberculosis prophage-like element,phiRv1

The genomes of Mycobacterium tuberculosis H37Rv and CDC1551 each contain two prophage-like elements, phiRv1 and phiRv2. The phiRv1 element is not only absent from Mycobacterium bovis BCG but is in different locations within the two sequenced M. tuberculosis genomes; in both cases phiRv1 is inserted into a REP13E12 repeated sequence, which presumably contains the bacterial attachment site, attB, for phiRv1. Although phiRv1 is probably too small to encode infectious phage particles, it may nevertheless have an active integration/excision system and be capable of moving from one chromosomal position to another. We show here that the M. tuberculosis H37Rv phiRv1 element does indeed encode an active site-specific recombination system in which an integrase of the serine recombinase family (Rv1586c) catalyses integration and excision and a small, basic phiRv1-encoded protein (Rv1584c) controls the directionality of re-combination. Integration-proficient plasmid vectors derived from phiRv1 efficiently transform BCG, can utilize four of the seven REP13E12 sites present in BCG as attachment sites, and can occupy more than one site simultaneously.     

Genetic analysis of peptidoglycan biosynthesis in mycobacteria: characterization of a ddlA mutant of Mycobacterium smegmatis

A temperature-sensitive mutant of Mycobacterium smegmatis was characterized that contains a mutation in ddlA, the gene encoding D-alanine:D-alanine ligase. Enzymatic assays using recombinant proteins and D-cycloserine susceptibility indicate that the A365V mutation in the SMEG35 DdlA protein causes a reduction in enzymatic activity in vitro and in vivo.     

Survival Rate and Enzyme Activities ofLactobacillus acidophilusFollowing Frozen Storage

The ability of two strains of Lactobacillus acidophilus, CRL 640 and CRL 800, to survive and retain their biological activities under frozen storage was determined. Freezing and thawing, as well as frozen storage, damaged the cell membrane, rendering the microorganisms sensitive to sodium chloride and bile salts. Both lactic acid production and proteolytic activity were depressed after 21 days at -20 degreesC, whereas beta-galactosidase activity per cell unit was increased. Cell injury was partially overcome after repair in a salt-rich medium. Copyright 1998 Academic Press.     

Genome sequence, structural proteins, and capsid organization of the cyanophage Syn5: a "horned" bacteriophage of marine synechococcus

Marine Synechococcus spp and marine Prochlorococcus spp are numerically dominant photoautotrophs in the open oceans and contributors to the global carbon cycle. Syn5 is a short-tailed cyanophage isolated from the Sargasso Sea on Synechococcus strain WH8109. Syn5 has been grown in WH8109 to high titer in the laboratory and purified and concentrated retaining infectivity. Genome sequencing and annotation of Syn5 revealed that the linear genome is 46,214 bp with a 237 bp terminal direct repeat. Sixty-one open reading frames (ORFs) were identified. Based on genomic organization and sequence similarity to known protein sequences within GenBank, Syn5 shares features with T7-like phages. The presence of a putative integrase suggests access to a temperate life cycle. Assignment of 11 ORFs to structural proteins found within the phage virion was confirmed by mass-spectrometry and N-terminal sequencing. Eight of these identified structural proteins exhibited amino acid sequence similarity to enteric phage proteins. The remaining three virion proteins did not resemble any known phage sequences in GenBank as of August 2006. Cryo-electron micrographs of purified Syn5 virions revealed that the capsid has a single “horn”, a novel fibrous structure protruding from the opposing end of the capsid from the tail of the virion. The tail appendage displayed an apparent 3-fold rather than 6-fold symmetry. An 18 Å resolution icosahedral reconstruction of the capsid revealed a T = 7 lattice, but with an unusual pattern of surface knobs. This phage/host system should allow detailed investigation of the physiology and biochemistry of phage propagation in marine photosynthetic bacteria.