A common catalytic mechanism for proteins of the HutI family

Imidazolonepropionase (HutI) (imidazolone-5-propanote hydrolase, EC 3.5.2.7) is a member of the amidohydrolase superfamily and catalyzes the conversion of imidazolone-5-propanoate to N-formimino-L-glutamate in the histidine degradation pathway. We have determined the three-dimensional crystal structures of HutI from Agrobacterium tumefaciens (At-HutI) and an environmental sample from the Sargasso Sea Ocean Going Survey (Es-HutI) bound to the product [ N-formimino-L-glutamate (NIG)] and an inhibitor [3-(2,5-dioxoimidazolidin-4-yl)propionic acid (DIP)], respectively. In both structures, the active site is contained within each monomer, and its organization displays the landmark feature of the amidohydrolase superfamily, showing a metal ligand (iron), four histidines, and one aspartic acid. A catalytic mechanism involving His265 is proposed on the basis of the inhibitor-bound structure. This mechanism is applicable to all HutI forms.     

Brca2 (XRCC11) Deficiency Results in Radioresistant DNA Synthesis and a Higher Frequency of Spontaneous Deletions

We show here that the radiosensitive Chinese hamster cell mutant (V-C8) of group XRCC11 is defective in the breast cancer susceptibility gene Brca2. The very complex phenotype of V-C8 cells is complemented by a single human chromosome 13 providing the BRCA2 gene, as well as by the murine Brca2 gene. The Brca2 deficiency in V-C8 cells causes hypersensitivity to various DNA-damaging agents with an extreme sensitivity toward interstrand DNA cross-linking agents. Furthermore, V-C8 cells show radioresistant DNA synthesis after ionizing radiation, suggesting that Brca2 deficiency affects cell cycle checkpoint regulation. In addition, V-C8 cells display tremendous chromosomal instability and a high frequency of abnormal centrosomes. The mutation spectrum at the hprt locus showed that the majority of spontaneous mutations in V-C8 cells are deletions, in contrast to wild-type V79 cells. A mechanistic explanation for the genome instability phenotype of Brca2-deficient cells is provided by the observation that the nuclear localization of the central DNA repair protein in homologous recombination, Rad51, is reduced in V-C8 cells.     

Tannase activity from the marine cyanobacterium Phormidium valderianum BDU140441

The marine cyanobacterium Phormidium valderianum BDU 140441 exhibited the ability to grow at 0.25?mM tannic acid, a known hindering chemical for microbial growth. The tannic acid-degrading ability of the organism is evident from the UV–visible absorption spectrum. In addition, the existence of tannase has been localized by activity staining, and its induction in activity upon tannic acid exposure was confirmed in native gel. The critical tannic acid metabolization enzymes tested for are polyphenol oxidase and esterases; both are well known for tannic acid degradation. Upon tannic acid exposure, increased activity of polyphenol oxidase and expression of few new isoforms of esterase were identified by activity staining.     

Isolation and identification of cytokinins from Euglena gracilis transfer ribonucleic acid.

Three ribonucleosides responsible for cytokinin activity in Euglena gracilis var Bacillaris tRNA have been isolated and identified as 6-(3-methyl-2-butenylamino)-9-beta-D-ribofuranosylpurine, 6-(4-hydroxy-3-methyl-cis-2-butenylamino)-9-beta-D-ribofuranosylpurine, and 6-(4-hydroxy-3-methyl-2-butenylamino)-2-methylthio-9-beta-D-ribofuranosylpurine. The structures of these compounds were assigned on the basis of their chromatographic properties and ultraviolet and mass spectra which were identical with those of the corresponding synthetic compounds. The elution profiles of cytokinin bioassay activity and of 35S radioactivity suggest the presence of a trace amount of 6-(3-methyl-2-butenylamino)-2-methylthio-9-beta-D-ribofuranosylpurine.     

Solution structure of 2',5' d(G4C4). Relevance to topological restrictions and nature's choice of phosphodiester links.

The NMR structure of 2',5' d(GGGGCCCC) was determined to gain insights into the structural differences between 2',5'- and 3',5'-linked DNA duplexes that may be relevant in elucidating nature's choice of sugar-phosphate links to encode genetic information. The oligomer assumes a duplex with extended nucleotide repeats formed out of mostly N-type sugar puckers. With the exception of the 5'-terminal guanine that assumes the syn glycosyl conformation, all other bases prefer the anti glycosyl conformation. Base pairs in the duplex exhibit slide (-1.96 A) and intermediate values for X-displacement (-3.23 A), as in ADNA, while their inclination to the helical axis is not prominent. Major and minor grooves display features intermediate to A and BDNA. The duplex structure of iso d(GGGGCCCC) may therefore be best characterized as a hybrid of A and BDNA. Importantly, the results confirm that even 3' deoxy 2',5' DNA supports duplex formation only in the presence of distinct slide (>or=-1.6 A) and X-displacement (>or=-2.5 A) for base pairs, and hence does not favor an ideal BDNA topology characterized by their near-zero values. Such restrictions on base pair movements in 2',5' DNA, which are clearly absent in 3',5' DNA, are expected to impose constraints on its ability for deformability of the kind observed in DNA during its compaction and interaction with proteins. It is therefore conceivable that selection pressure relating to the optimization of topological features might have been a factor in the rejection of 2',5' links in preference to 3',5' links.