Oxford Solutions Intermediate
Dodie Clymore
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The cryoenzymology of several different beta-lactamases has been investigated. Particular attention has been paid to the experimental pitfalls of the technique. These include such factors as false bursts at the start of the reaction, instability of the enzymes during turnover, and Km values so high that little of the enzyme is present as a complex. Many of the difficulties in cryoenzymology stem from the use of organic cryosolvents. A novel "salt" cryosolvent has been tested: ammonium acetate solutions can be used down to about -60 degrees C. The enzymes examined are readily soluble, and stable, in this solvent. Nevertheless, out of 17 beta-lactamase beta-lactam systems, only 4 proved suitable for detailed investigation. In two of these, the hydrolysis of nitrocefin or 7-(thienyl-2-acetamido)-3-[[2-[[4- (dimethylamino)phenyl]azo]pyridinio]-methyl]cephem-4-carboxylic acid (PADAC), by beta-lactamase I from Bacillus cereus, substrate was converted into product at a slow enough rate (at -60 or -55 degrees C, respectively) for it to be possible to do successive scans during the course of the reaction. The spectra were those of substrate and product, and no intermediate was detected. The results argue against the accumulation of intermediate acyl-enzyme. The hydrolysis of PADAC by the P99 beta-lactamase from Enterobacter cloacae again showed spectra characteristic of substrate and product, and there was, moreover, a break in the Arrhenius plot; it is possible that a conformational change is (at least partially) rate-determining. The hydrolysis of dinitrophenylpenicillin by the P99 beta-lactamase did show features suggesting the accumulation of acyl-enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)
N2 - In recent years, collaborative multi-stakeholder initiatives (MSIs) between private sector companies and civil society organizations have sought to find common solutions to sustainability challenges related to cotton, timber, and other raw materials, as well as hazardous work conditions in industries such as garments, textiles, and leather production. In this article, we contribute to the MSI literature by conceptualizing how intermediate standard implementing organizations deal with organizational tensions as street level bureaucrats, theorizing their differential capacity to navigate between two sets of competing pressures inherent to MSI standards: (1) upscaling in volume terms and maintaining the stringency of the standard; and (2) farmer capacity building and auditing approaches. Empirically, we analyze how intermediary standard implementing organizations of a particular MSI, the Better Cotton Initiative (BCI), mediate between these global sustainability standards pressures and the needs of farmers in Pakistan and India. We conclude that this process of mediating between global standards and local farmer needs results in a situation whereby BCI intermediate standard implementing organizations increasingly spend time on data gathering exercises instead of enabling farmers to comply with the standard through capacity building.
AB - In recent years, collaborative multi-stakeholder initiatives (MSIs) between private sector companies and civil society organizations have sought to find common solutions to sustainability challenges related to cotton, timber, and other raw materials, as well as hazardous work conditions in industries such as garments, textiles, and leather production. In this article, we contribute to the MSI literature by conceptualizing how intermediate standard implementing organizations deal with organizational tensions as street level bureaucrats, theorizing their differential capacity to navigate between two sets of competing pressures inherent to MSI standards: (1) upscaling in volume terms and maintaining the stringency of the standard; and (2) farmer capacity building and auditing approaches. Empirically, we analyze how intermediary standard implementing organizations of a particular MSI, the Better Cotton Initiative (BCI), mediate between these global sustainability standards pressures and the needs of farmers in Pakistan and India. We conclude that this process of mediating between global standards and local farmer needs results in a situation whereby BCI intermediate standard implementing organizations increasingly spend time on data gathering exercises instead of enabling farmers to comply with the standard through capacity building.
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