Can ultrasonic energy efficiently speed (18)O-labeling of proteins?,
Carreira, Ricardo J., Lodeiro Carlos, Diniz Mario S., Moura Isabel, and Capelo Jose L.
, Proteomics, Nov, Volume 9, Number 21, p.4974-4977, (2009)
AbstractWe report in this work on the robustness of ultrasonic energy as a tool to speed the isotopic labeling of proteins using the (18)O-decoupling procedure. The first part of the decoupling procedure, comprising protein denaturation, reduction, alkylation and digestion, is done in 8 min under the effects of an ultrasonic field whilst the second part, the isotopic labeling, was assayed with and without the use of ultrasonic energy. Our results clearly demonstrate that the (18)O-isotopic labeling in a decoupling procedure cannot be accelerated using an ultrasonic field.
Can ultrasonic energy efficiently speed (18)O-labeling of proteins?,
Carreira, Ricardo J., Lodeiro Carlos, Diniz Mario S., Moura Isabel, and Capelo Jose L.
, Proteomics, Nov, Volume 9, Number 21, p.4974-4977, (2009)
AbstractWe report in this work on the robustness of ultrasonic energy as a tool to speed the isotopic labeling of proteins using the (18)O-decoupling procedure. The first part of the decoupling procedure, comprising protein denaturation, reduction, alkylation and digestion, is done in 8 min under the effects of an ultrasonic field whilst the second part, the isotopic labeling, was assayed with and without the use of ultrasonic energy. Our results clearly demonstrate that the (18)O-isotopic labeling in a decoupling procedure cannot be accelerated using an ultrasonic field.
A novel nitrite biosensor based on conductometric electrode modified with cytochrome c nitrite reductase composite membrane,
Zhang, Z., Xia S., Leonard D., Jaffrezic-Renault N., Zhang J., Bessueille F., Goepfert Y., Wang X., Chen L., Zhu Z., Zhao J., Almeida M. G., and Silveira C. M.
, Biosensors & Bioelectronics, Feb 15, Volume 24, Number 6, p.1574-9, (2009)
AbstractA conductometric biosensor for nitrite detection was developed using cytochrome c nitrite reductase (ccNiR) extracted from Desulfovibrio desulfuricans ATCC 27774 cells immobilized on a planar interdigitated electrode by cross-linking with saturated glutaraldehyde (GA) vapour in the presence of bovine serum albumin, methyl viologen (MV), Nafion, and glycerol. The configuration parameters for this biosensor, including the enzyme concentration, ccNiR/BSA ratio, MV concentration, and Nafion concentration, were optimized. Various experimental parameters, such as sodium dithionite added, working buffer solution, and temperature, were investigated with regard to their effect on the conductance response of the biosensor to nitrite. Under the optimum conditions at room temperature (about 25 degrees C), the conductometric biosensor showed a fast response to nitrite (about 10s) with a linear range of 0.2-120 microM, a sensitivity of 0.194 microS/microM [NO(2)(-)], and a detection limit of 0.05 microM. The biosensor also showed satisfactory reproducibility (relative standard deviation of 6%, n=5). The apparent Michaelis-Menten constant (K(M,app)) was 338 microM. When stored in potassium phosphate buffer (100mM, pH 7.6) at 4 degrees C, the biosensor showed good stability over 1 month. No obvious interference from other ionic species familiar in natural waters was detected. The application experiments show that the biosensor is suitable for use in real water samples.
Molybdenum induces the expression of a protein containing a new heterometallic Mo-Fe cluster in Desulfovibrio alaskensis,
Rivas, M. G., Carepo M. S., Mota C. S., Korbas M., Durand M. C., Lopes A. T., Brondino C. D., Pereira A. S., George G. N., Dolla A., Moura J. J., and Moura I.
, Biochemistry, Feb 10, Volume 48, Number 5, p.873-82, (2009)
AbstractThe characterization of a novel Mo-Fe protein (MorP) associated with a system that responds to Mo in Desulfovibrio alaskensis is reported. Biochemical characterization shows that MorP is a periplasmic homomultimer of high molecular weight (260 +/- 13 kDa) consisting of 16-18 monomers of 15321.1 +/- 0.5 Da. The UV/visible absorption spectrum of the as-isolated protein shows absorption peaks around 280, 320, and 570 nm with extinction coefficients of 18700, 12800, and 5000 M(-1) cm(-1), respectively. Metal content, EXAFS data and DFT calculations support the presence of a Mo-2S-[2Fe-2S]-2S-Mo cluster never reported before. Analysis of the available genomes from Desulfovibrio species shows that the MorP encoding gene is located downstream of a sensor and a regulator gene. This type of gene arrangement, called two component system, is used by the cell to regulate diverse physiological processes in response to changes in environmental conditions. Increase of both gene expression and protein production was observed when cells were cultured in the presence of 45 microM molybdenum. Involvement of this system in Mo tolerance of sulfate reducing bacteria is proposed.