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Maxim Blum - One of the best experts on this subject based on the ideXlab platform.

Santosh O. Ramchuran - One of the best experts on this subject based on the ideXlab platform.

  • The methylotrophic yeast Pichia pastoris as a host for the expression and production of thermostable xylanase from the bacterium Rhodothermus marinus
    FEMS Yeast Research, 2005
    Co-Authors: Santosh O. Ramchuran, Bruno Mateus, Olle Holst, Eva Nordberg Karlsson
    Abstract:

    A thermostable glycoside hydrolase family-10 xylanase originating from Rhodothermus marinus was cloned and expressed in the methylotrophic yeast Pichia pastoris (SMD1168H). The DNA sequence from Rmxyn10A encoding the xylanase catalytic module was PCR-amplified and cloned in frame with the Saccharomyces cerevisiae alpha-factor secretion signal under the control of the alcohol oxidase (AOX1) promotor. Optimisation of enzyme production in batch fermentors, with methanol as a sole carbon source, enabled secretion yields up to 3gl(-1) xylanase with a maximum activity of 3130Ul(-1) to be achieved. N-terminal sequence analysis of the heterologous xylanase indicated that the secretion signal was correctly processed in P. pastoris and the molecular weight of 37kDa was in agreement with the theoretically calculated molecular Mass. Introduction of a heat-pretreatment step was however necessary in order to fold the heterologous xylanase to an active state, and at the conditions used this step yielded a 200-fold increase in xylanase activity. Thermostability of the produced xylanase was monitored by differential-scanning calorimetry, and the transition temperature (T(m)) was 78 degrees C. R. marinus xylanase is the first reported thermostable gram-negative bacterial xylanase efficiently secreted by P. pastoris.

  • The methylotrophic yeast Pichia pastoris as a host for the expression and production of thermostable xylanase from the bacterium
    2005
    Co-Authors: Santosh O. Ramchuran, Bruno Mateus, Olle Holst, Eva Nordberg
    Abstract:

    A thermostable glycoside hydrolase family 10 xylanase originating from Rhodothermus marinus was cloned and expressed in the methylotrophic yeast Pichia pastoris (SMD1168H). The DNA sequence from Rmxyn 10A encoding the xylanase catalytic module was PCR-amplified and cloned in frame with the Saccharomyces cerevisiae α-factor secretion signal under the control of the alcohol oxidase (AOX1) promotor. Optimisation of enzyme production in batch fermentors with methanol as a sole carbon source, enabled secretion yields up to 3 grams xylanase per litre with a maximum activity of 3130 U/L to be achieved. N-terminal sequence analysis of the heterologous xylanase indicated that the secretion signal was correctly processed in P. pastoris and the molecular weight of 37 kDa was in agreement with the theoretically calculated molecular Mass. Introduction of a heat-pretreatment step was however necessary in order to fold the heterologous xylanase to an active state, and at the conditions used this step yielded a 200-fold increase in xylanase activity. Thermostability of the produced xylanase was monitored by differential scanning calorimetry, and the transition temperature (Tm) was 78 °C. R. marinus xylanase is the first reported thermostable gram-negative bacterial xylanase efficiently secreted

Raul F Velazquez - One of the best experts on this subject based on the ideXlab platform.

Kenneth J Mcleod - One of the best experts on this subject based on the ideXlab platform.

  • prevention of postmenopausal bone loss by a low magnitude high frequency mechanical stimuli a clinical trial assessing compliance efficacy and safety
    Journal of Bone and Mineral Research, 2003
    Co-Authors: Clinton T Rubin, Robert R Recker, D M Cullen, John P Ryaby, Joan M Mccabe, Kenneth J Mcleod
    Abstract:

    A 1-year prospective, randomized, double-blind, and placebo-controlled trial of 70 postmenopausal women demonstrated that brief periods (<20 minutes) of a low-level (0.2g, 30 Hz) vibration applied during quiet standing can effectively inhibit bone loss in the spine and femur, with efficacy increasing significantly with greater compliance, particularly in those subjects with lower body Mass. Introduction: Indicative of the anabolic potential of mechanical stimuli, animal models have demonstrated that short periods (<30 minutes) of low-magnitude vibration (<0.3g), applied at a relatively high frequency (20–90 Hz), will increase the number and width of trabeculae, as well as enhance stiffness and strength of cancellous bone. Here, a 1-year prospective, randomized, double-blind, and placebo-controlled clinical trial in 70 women, 3–8 years past the menopause, examined the ability of such high-frequency, low-magnitude mechanical signals to inhibit bone loss in the human. Materials and Methods: Each day, one-half of the subjects were exposed to short-duration (two 10-minute treatments/day), low-magnitude (2.0 m/s2 peak to peak), 30-Hz vertical accelerations (vibration), whereas the other half stood for the same duration on placebo devices. DXA was used to measure BMD at the spine, hip, and distal radius at baseline, and 3, 6, and 12 months. Fifty-six women completed the 1-year treatment. Results and Conclusions: The detection threshold of the study design failed to show any changes in bone density using an intention-to-treat analysis for either the placebo or treatment group. Regression analysis on the a priori study group demonstrated a significant effect of compliance on efficacy of the intervention, particularly at the lumbar spine (p = 0.004). Posthoc testing was used to assist in identifying various subgroups that may have benefited from this treatment modality. Evaluating those in the highest quartile of compliance (86% compliant), placebo subjects lost 2.13% in the femoral neck over 1 year, whereas treatment was associated with a gain of 0.04%, reflecting a 2.17% relative benefit of treatment (p = 0.06). In the spine, the 1.6% decrease observed over 1 year in the placebo group was reduced to a 0.10% loss in the active group, indicating a 1.5% relative benefit of treatment (p = 0.09). Considering the interdependence of weight, the spine of lighter women (<65 kg), who were in the highest quartile of compliance, exhibited a relative benefit of active treatment of 3.35% greater BMD over 1 year (p = 0.009); for the mean compliance group, a 2.73% relative benefit in BMD was found (p = 0.02). These preliminary results indicate the potential for a noninvasive, mechanically mediated intervention for osteoporosis. This non-pharmacologic approach represents a physiologically based means of inhibiting the decline in BMD that follows menopause, perhaps most effectively in the spine of lighter women who are in the greatest need of intervention.

Eva Nordberg - One of the best experts on this subject based on the ideXlab platform.

  • The methylotrophic yeast Pichia pastoris as a host for the expression and production of thermostable xylanase from the bacterium
    2005
    Co-Authors: Santosh O. Ramchuran, Bruno Mateus, Olle Holst, Eva Nordberg
    Abstract:

    A thermostable glycoside hydrolase family 10 xylanase originating from Rhodothermus marinus was cloned and expressed in the methylotrophic yeast Pichia pastoris (SMD1168H). The DNA sequence from Rmxyn 10A encoding the xylanase catalytic module was PCR-amplified and cloned in frame with the Saccharomyces cerevisiae α-factor secretion signal under the control of the alcohol oxidase (AOX1) promotor. Optimisation of enzyme production in batch fermentors with methanol as a sole carbon source, enabled secretion yields up to 3 grams xylanase per litre with a maximum activity of 3130 U/L to be achieved. N-terminal sequence analysis of the heterologous xylanase indicated that the secretion signal was correctly processed in P. pastoris and the molecular weight of 37 kDa was in agreement with the theoretically calculated molecular Mass. Introduction of a heat-pretreatment step was however necessary in order to fold the heterologous xylanase to an active state, and at the conditions used this step yielded a 200-fold increase in xylanase activity. Thermostability of the produced xylanase was monitored by differential scanning calorimetry, and the transition temperature (Tm) was 78 °C. R. marinus xylanase is the first reported thermostable gram-negative bacterial xylanase efficiently secreted