The UvrA, UvrB and UvrC proteins of E. coli are subunits of a DNA repair enzyme, the ABC exonuclease. In this paper we study the uvrC regulatory region. The uvrC structural gene is preceded by an open reading frame encoding a 24 kD protein. A uvrC promoter has been mapped within this gene. The transcription start of a second promoter located 5' of the 24 kD gene is mapped in vivo. We show that transcription from both promoters on the chromosome is not inducible by UV damage. The possible translation start codons of the UvrC and of the 24 kD protein are determined. Sequences encoding the N-terminal part of the UvrC protein overlap with sequences encoding the C-terminal part of the 24 kD protein. To examine a possible function of the 24 kD gene in repair, a 24 kD insertion mutant was created in the chromosome. The mutant however only slightly affects the UV sensitivity of the cell. Transcription of P3 alone provides sufficient UvrC protein for the normal repair of UV lesions.

Several robotic exploration missions will travel to Mars during this decade to investigate habitability and the possible presence of life. Field research at Mars analogue sites such as desert environments can provide important constraints for instrument calibration, landing site strategies and expected life detection targets. We have characterized the mineralogy, organic chemistry and microbiology of ten selected sample sites from the Utah desert in close vicinity to the Mars Desert Research Station (MDRS) during the EuroGeoMars 2009 campaign (organized by International Lunar Exploration Working Group (ILEWG), NASA Ames and ESA ESTEC). Compared with extremely arid deserts (such as the Atacama), organic and biological materials can be identified in a larger number of samples and subsequently be used to perform correlation studies. Among the important findings of this field research campaign are the diversity in the mineralogical composition of soil samples even when collected in close proximity, the low abundances of detectable polycyclic aromatic hydrocarbons (PAHs) and amino acids and the presence of biota of all three domains of life with significant heterogeneity. An extraordinary variety of putative extremophiles, mainly Bacteria and also Archaea and Eukarya was observed. The dominant factor in measurable bacterial abundance seems to be soil porosity and lower small (clay-sized) particle content. However, correlations between many measured parameters are difficult to establish. Field research conducted during the EuroGeoMars 2009 campaign shows that the geological history and depositional environment of the region, as well as the mineralogy influence the ability to detect compounds such as amino acids and DNA. Clays are known to strongly absorb and bind organic molecules often preventing extraction by even sophisticated laboratory methods. Our results indicate the need for further development and optimization of extraction procedures that release biological compounds from host matrices to enable the effective detection of biomarkers during future sampling campaigns on Earth and Mars.

Preferential repair of UV-induced damage is a phenomenon by which mammalian cells might enhance their survival. This paper presents the first evidence that preferential repair occurs in the lower eukaryote Saccharomyces cerevisiae. Moreover an unique approach is reported to compare identical sequences present on the same chromosome and only differing in expression. We determined the removal of pyrimidine dimers from two identical alpha-mating type loci and we were able to show that the active MAT alpha locus is repaired preferentially to the inactive HML alpha locus. In a sir-3 mutant, in which both loci are active this preference is not observed.

Bacteriophage Ab2ate [Ab2cI857intam6(AbioAB)bioFCD'uvrB+phr'] codes for a function(s) that confers UV resistance (Uvr+) and reactivation of irradiated phage (Hcr+) to an Uvr-Hcr-Escherichia coli strain. It was demonstrated that these functions are expressed under the control of bacterial regulatory elements located on Xb2att2 DNA. The location of the E. coli uvrB gene on the DNA of this transducing phage was established by heteroduplex and restriction-enzyme analyses. Recombinant DNA molecules were constructed in vitro from plasmid pMB9 (Tcr), as the vector, and an EcoRI fragment (EcoRI-F) of Xb2att2 DNA. The resulting plasmid, designated pNP5, has a molecular weight of 5.1 x 106 and replicates in a relaxed fashion. Transformation of E. coli uvrB with plasmid pNP5 resulted in clones that are Uvr+ Tcr. Irradiation of bacteria transformed with plasmid pNP5 with low UV doses revealed a complete restoration of the Uvr+ phenotype by the presence of the cloned EcoRI-F DNA, while only a partial restoration was observed after irradiation with high UV doses. Likewise, the Hcr+ character was also partially restored due to the presence of pNP5. No correlation was found between the acquired Uvr+, Hcr+ properties, and the presence of correndonuclease II activity in an extract of bacteria that harbor plasmid pNP5. This work HP3404 KMBL1001(pNP5 Tcr)Uvr+ This work a All bacterial strains are F- .

The gene encoding quinohacmoprotein ethanol dehydrogenase type I (QH-EDH,) from Cornamonas testnsteroizi has been cloned and sequenced. Coniparjson of thc umino acid sequence deduced from this with that determined for the N-terminal amino acid strelch of purified QH-EDH,, suggests that the gene also contains a leadcr sequence of 31 residues. Based on this information, the rnolccular mass of the apoenzyme, i.e. thc enzyme without the cofactors pyrroloquinoline quinone (PQQ) and haem c, and without the C d ' , appears to be 73 200 Da. Alignment of the deduced amino acid sequence to that of other PQQcontaining dehydrogenascs showed that good similarity (up to 43% identity) exists with most of them. This also showed (hat the amino acid residues presumed to he involved in PQQ and Ca2+ binding and in the typical features of structure and catalysis of inethanol dehydrogcnase, are conservcd at the same positions in QH-EDH,. The C-terminal part of the protcin, containing the haem c, exhibited some sirnilarity to cytochromes us,, from cyanobacteria and algae. Correct processing of the yhedh gene appeared lo occur in Escherichio coti stmin JM 109 in which the gene was placed under control of the Iuc promoter, iis judged from a positive reaction with antibodies raised against authentic QH-EDH,, the size of the protcin, the prescnce of haem c in it, and the spccific activity value obtained after reconstitution with PQQ. The yhedh gene seems to form part of an operon which is organized in a way different from that of the genes required for methanol oxidation in methylotrophic bacteria.Kqwords: quinoprotein; pyrroloquinoline quinone: alcohol dehydrogenase; heme c'; Coinanlonus testosteroni.Oxidation of alcohols to aldehydes can proceed via catalysis by oxidases or dehydrogenascs. The dehydrogenases can be subdivided into the NAD(P)-depcnctent and the dye-linked ones. It has been found for many of thc dye-linked alcohol dehydrogenases occurring in Gram-negative bacteria that PQQ [pyrroloquinoline quinone, i.e. 2,7.9-triciirboxy-1 H-pyrrolo(2,3,f)quinoline-4,s-dionej functions as cofactor [ 1, 21.Two groups of PQQ-dependent alcohol dehydrogenases can be distinguished, the quinoprolein alcohol dehydrogenases (containing PQQ and Caz' as non-proteinaccous compounds) and the quinohaemoprotein alcohol dehydrogenases (containing PQQ, CaZ' as well as haem c').The group of quinoprotein alcohol dchydrogenases comprises methanol dehydrogenases (MDH), occurring i n rnethyl- 121 and is induced when thc bacterium is grown on rnedia with ethuuol (or butanol) as sole source ol carbon and energy. Curiously, the cnzyme is produced in its apo-form, lacking the cofiictor PQQ. Easy reconstitution to active holo-enzyme occurs with PQQ in the presence of Cii" , in vivo as well BS in vitro. Indications have becn obtained that the apo-enzyme contains a sensitivc spot for proteolytic attack in the stationary growth phase,