Evolution of Protein Molecules

The selective transformation of ubiquitous but inert C–H bonds to other functional groups has far-reaching practical implications, ranging from more efficient strategies for fine chemical synthesis to the replacement of current petrochemical feedstocks by less expensive and more readily available alkanes. The past twenty years have seen many examples of C–H bond activation at transition-metal centres, often under remarkably mild conditions and with high selectivity. Although profitable practical applications have not yet been developed, our understanding of how these organometallic reactions occur, and what their inherent advantages and limitations for practical alkane conversion are, has progressed considerably. In fact, the recent development of promising catalytic systems highlights the potential of organometallic chemistry for useful C–H bond activation strategies that will ultimately allow us to exploit Earth's alkane resources more efficiently and cleanly.

Understanding and exploiting C–H bond activation

The use of biocatalysis for industrial synthetic chemistry is on the verge of significant growth. Biocatalytic processes can now be carried out in organic solvents as well as aqueous environments, so that apolar organic compounds as well as water-soluble compounds can be modified selectively and efficiently with enzymes and biocatalytically active cells. As the use of biocatalysis for industrial chemical synthesis becomes easier, several chemical companies have begun to increase significantly the number and sophistication of the biocatalytic processes used in their synthesis operations.

Industrial biocatalysis today and tomorrow

Recent advances in molecular biology have extended our understanding of the metabolic processes related to microbial transformation of petroleum hydrocarbons. The physiological responses of microorganisms to the presence of hydrocarbons, including cell surface alterations and adaptive mechanisms for uptake and efflux of these substrates, have been characterized. New molecular techniques have enhanced our ability to investigate the dynamics of microbial communities in petroleum-impacted ecosystems. By establishing conditions which maximize rates and extents of microbial growth, hydrocarbon access, and transformation, highly accelerated and bioreactor-based petroleum waste degradation processes have been implemented. Biofilters capable of removing and biodegrading volatile petroleum contaminants in air streams with short substrate-microbe contact times ( 2 S and sulfoxides from petrochemical waste streams. Microbes also have potential for use in removal of nitrogen from crude oil leading to reduced nitric oxide emissions provided that technical problems similar to those experienced in biodesulfurization can be solved. Enzymes are being exploited to produce added-value products from petroleum substrates, and bacterial biosensors are being used to analyze petroleum-contaminated environments.

Recent Advances in Petroleum Microbiology

http://bioprocesos.unq.edu.ar/Biopro%20II/Paper%20escalado.pdf

Bioreactor scale-up and oxygen transfer rate in microbial processes: An overview

Miniaturized growth systems for heterogeneous culture collections are not only attractive in reducing demands for incubation space and medium but also in making the parallel handling of large numbers of strains more practicable. We report here on the optimization of oxygen transfer rates in deep-well microtiter plates and the development of a replication system allowing the simultaneous and reproducible sampling of 96 frozen glycerol stock cultures while the remaining culture volume remains frozen. Oxygen transfer rates were derived from growth curves of Pseudomonas putida and from rates of oxygen disappearance due to the cobalt-catalyzed oxidation of sulfite. Maximum oxygen transfer rates (38 mmol liter 21 h 21 , corresponding to a mass transfer coefficient of 188 h 21 ) were measured during orbital shaking at 300 rpm at a shaking diameter of 5 cm and a culture volume of 0.5 ml. A shaking diameter of 2.5 cm resulted in threefold-lower values. These high oxygen transfer rates allowed P. putida to reach a cell density of approximately 9 g (dry weight) liter 21 during growth on a glucose mineral medium at culture volumes of up to 1 ml. The growth-and-replication system was evaluated for a culture collection consisting of aerobic strains, mainly from the genera Pseudomonas, Rhodococcus, and Alcaligenes, using mineral media and rich media. Cross-contamination and excessive evaporation during vigorous aeration were adequately prevented by the use of a sandwich cover of spongy silicone and cotton wool on top of the microtiter plates.

/pdf/methods-for-intense-aeration-growth-storage-and-replication-huvvg9me91.pdf

Methods for Intense Aeration, Growth, Storage, and Replication of Bacterial Strains in Microtiter Plates

The first step in the aerobic degradation of alkanes by bacteria, yeasts, and fungi is catalyzed by oxygenases. These enzymes, which introduce oxygen atoms derived from molecular oxygen into the alkane substrate, play an important role in oil bioremediation and in the cometabolic degradation of compounds such as trichloroethylene and fuel oxygenates. In addition, they are useful biocatalysts and important models for a difficult chemical reaction: the regio- and stereospecific activation of C-H bonds. Several unrelated enzyme classes catalyze the oxidation of alkanes. Alkane-degrading yeast strains contain multiple alkane hydroxylases belonging to the P450 superfamily, while many bacteria contain enzymes related to the Pseudomonas putida GPo1 membrane-bound alkane hydroxylase system. Short-chain alkanes are probably oxidized by alkane hydroxylases related to the soluble and particulate methane monooxygenases. Only the membrane-bound enzymes have been studied with respect to their prevalence in environments such as soils or aquifers.

/pdf/diversity-of-alkane-hydroxylase-systems-in-the-environment-3sunggxnqh.pdf

Diversity of Alkane Hydroxylase Systems in the Environment

https://www.cell.com/trends/biotechnology/pdf/S0167-7799(03)00032-5.pdf

Practical issues in the application of oxygenases

Using proteins in their natural environment: potential and limitations of microbial whole-cell hydroxylations in applied biocatalysis.

The past 5 years have seen significant progress in the field of limonene biotransformation, especially with regard to the regiospecificity of microbial biocatalysts. Whereas earlier only regiospecific biocatalysts for the 1,2 position (limonene-1,2-diol) and the 8-position (α-terpineol) were available, recent reports describe microbial biocatalysts specifically hydroxylating the 3-position (isopiperitenol), 6-position (carveol and carvone), and 7-position (perillyl alcohol, perillylaaldehyde, and perillic acid). The present review also includes the considerable progress made in the characterization of plant P-450 limonene hydroxylases and the cloning of the encoding genes.

Wouter A. Duetz

Papers

Methods for Intense Aeration, Growth, Storage, and Replication of Bacterial Strains in Microtiter Plates

Diversity of Alkane Hydroxylase Systems in the Environment

Practical issues in the application of oxygenases

Using proteins in their natural environment: potential and limitations of microbial whole-cell hydroxylations in applied biocatalysis.

Biotransformation of limonene by bacteria, fungi, yeasts, and plants