Extremely halophilic pleomorphic archaeal virus HRPV9 extends the diversity of pleolipoviruses with integrases.

Certain pleomorphic archaeal viruses are highly infectious even at saturated salt. These viruses belong to the genus Betapleolipovirus of the recently described archaeal virus family Pleolipoviridae. Pleolipoviruses comprise single-stranded or double-stranded, circular or linear DNA genomes that share countless homologues among various archaeal genetic elements. Here we describe a new extremely halophilic betapleolipovirus, Halorubrum pleomorphic virus 9 (HRPV9), which has an integrase gene. We also identified new genes encoding minor pleolipoviral structural proteins. The studies on HRPV9 enhance our knowledge on pleolipoviruses, especially their reciprocal relatedness and relation to certain archaeal plasmids, proviruses and membrane vesicles.

The structure of the NTPase that powers DNA packaging into Sulfolobus turreted icosahedral virus 2.

Biochemical reactions powered by ATP hydrolysis are fundamental for the movement of molecules and cellular structures. One such reaction is the encapsidation of the double-stranded DNA (dsDNA) genome of an icosahedrally symmetric virus into a preformed procapsid with the help of a genome-translocating NTPase. Such NTPases have been characterized in detail from both RNA and tailed DNA viruses. We present four crystal structures and the biochemical activity of a thermophilic NTPase, B204, from the nontailed, membrane-containing, hyperthermoacidophilic archaeal dsDNA virus Sulfolobus turreted icosahedral virus 2. These are the first structures of a genome-packaging NTPase from a nontailed, dsDNA virus with an archaeal host. The four structures highlight the catalytic cycle of B204, pinpointing the molecular movement between substrate-bound (open) and empty (closed) active sites. The protein is shown to bind both single-stranded and double-stranded nucleic acids and to have an optimum activity at 80°C and pH 4.5. The overall fold of B204 places it in the FtsK-HerA superfamily of P-loop ATPases, whose cellular and viral members have been suggested to share a DNA-translocating mechanism.

The structure of an archaeal viral integrase reveals an evolutionarily conserved catalytic core yet supports a mechanism of DNA cleavage in trans.

The first structure of a catalytic domain from a hyperthermophilic archaeal viral integrase reveals a minimal fold similar to that of bacterial HP1 integrase and defines structural elements conserved across three domains of life. However, structural superposition on bacterial Holliday junction complexes and similarities in the C-terminal tail with that of eukaryotic Flp suggest that the catalytic tyrosine and an additional active-site lysine are delivered to neighboring subunits in trans. An intramolecular disulfide bond contributes significant thermostability in vitro.

Metagenomics and recovery of enzyme genes from alkaline saline environments.

Enzymes functioning at alkaline pH are widely used in the detergent industry as additives to improve the stain removal properties of domestic and industrial cleaning products. This industry provides by far the major mass market for enzymes. With constantly changing formulations in detergents and concerns over energy demands, new and improved enzymes are constantly in demand. Soda lakes host dense populations of alkali-loving microbes and, as such, provide vast reservoirs of potentially useful enzymes for such an industry. Traditional recovery methods for new enzymes have involved the isolation of microbes, preferably from a compatible chemical environment such as a soda lake, followed by screening of the isolates for useful enzymic activity. At least two commercially significant enzymes originating from soda lake microbes have been marketed following this route. However, the failure to cultivate more than a small percentage of microbes from most environments necessarily markedly reduces the recovery of new enzymes. In recent years, interest has focussed on more comprehensive recovery methods based around detecting appropriate enzyme genes in nucleic acids extracted from potentially useful sites, thus maximizing coverage of the whole genetic resource in a particular biotope. Here we review progress to date in soda lake biotopes and discuss ways the field may develop in the future.

Purification, crystallization and preliminary X-ray diffraction studies of the archaeal virus resolvase SIRV2.

The Holliday junction (or four-way junction) is the universal DNA intermediate whose interaction with resolving proteins is one of the major events in the recombinational process. These proteins, called DNA junction-resolving enzymes or resolvases, bind to the junction and catalyse DNA cleavage, promoting the release of two DNA duplexes. SIRV2 Hjc, a viral resolvase infecting a thermophylic archaeon, has been cloned, expressed and purified. Crystals have been obtained in space group C2, with unit-cell parameters a = 147.8, b = 99.9, c = 87.6, beta = 109.46 degrees, and a full data set has been collected at 3.4 A resolution. The self-rotation function indicates the presence of two dimers in the asymmetric unit and a high solvent content (77%). Molecular-replacement trials using known similar resolvase structures have so far been unsuccessful, indicating possible significant structural rearrangements.