A Versatile Aldehyde: Ferredoxin Oxidoreductase from the Organic Acid Reducing Thermoanaerobacter sp. Strain X514.

Aldehyde:ferredoxin oxidoreductases (AORs) have been isolated and biochemically-characterized from a handful of anaerobic or facultative aerobic archaea and bacteria. They catalyze the ferredoxin (Fd)-dependent oxidation of aldehydes to acids. Recently, the involvement of AOR in the reduction of organic acids to alcohols with electrons derived from sugar or synthesis gas was demonstrated, with alcohol dehydrogenases (ADHs) carrying out the reduction of the aldehyde to the alcohol (AOR-ADH pathway). Here, we describe the biochemical characterization of an AOR of the thermophilic fermentative bacterium Thermoanaerobacter sp. strain X514 (AORX514). The putative aor gene (Teth514_1380) including a 6x-His-tag was introduced into the genome of the genetically-accessible, related species Thermoanaerobacter kivui. The protein was purified to apparent homogeneity, and indeed revealed AOR activity, as measured by acetaldehyde-dependent ferredoxin reduction. AORX514 was active over a wide temperature (10 to 95 °C) and pH (5.5 to 11.5) range, utilized a wide variety of aldehydes (short and branched-chained, aliphatic, aromatic) and resembles archaeal sensu stricto AORs, as the protein is active in a homodimeric form. The successful, recombinant production of AORX514 in a related, well-characterized and likewise strict anaerobe paves the road towards structure-function analyses of this enzyme and possibly similar oxygen-sensitive or W/Mo-dependent proteins in the future.

Alcohol dehydrogenases AdhE and AdhB with broad substrate ranges are important enzymes for organic acid reduction in Thermoanaerobacter sp. strain X514.

BACKGROUND: The industrial production of various alcohols from organic carbon compounds may be performed at high rates and with a low risk of contamination using thermophilic microorganisms as whole-cell catalysts. Thermoanaerobacter species that thrive around 50-75 °C not only perform fermentation of sugars to alcohols, but some also utilize different organic acids as electron acceptors, reducing them to their corresponding alcohols. RESULTS: We purified AdhE as the major NADH- and AdhB as the major NADPH-dependent alcohol dehydrogenase (ADH) from the cell extract of the organic acid-reducing Thermoanaerobacter sp. strain X514. Both enzymes were present in high amounts during growth on glucose with and without isobutyrate, had broad substrate spectra including different aldehydes, with high affinities (< 1 mM) for acetaldehyde and for NADH (AdhE) or NADPH (AdhB). Both enzymes were highly thermostable at the physiological temperature of alcohol production. In addition to AdhE and AdhB, we identified two abundant AdhA-type ADHs based on their genes, which were recombinantly produced and biochemically characterized. The other five ADHs encoded in the genome were only expressed at low levels. CONCLUSIONS: According to their biochemical and kinetic properties, AdhE and AdhB are most important for ethanol formation from sugar and reduction of organic acids to alcohols, while the role of the two AdhA-type enzymes is less clear. AdhE is the only abundant aldehyde dehydrogenase for the acetyl-CoA reduction to aldehydes, however, acid reduction may also proceed directly by aldehyde:ferredoxin oxidoreductase. The role of the latter in bio-alcohol formation from sugar and in organic acid reduction needs to be elucidated in future studies.

The emerging role of aldehyde:ferredoxin oxidoreductases in microbially-catalyzed alcohol production.

The development of a bio-refinery industry based on liquid fuels is presumably key to successful replacement of fossil fuels and a reduction of carbon dioxide (CO2) emissions. Ethanol and longer-chain alcohols are supposed to play a key role since they are relatively easy to produce, using microorganisms as whole-cell biocatalysts. Alcohols may be produced from lignocellulose-derived biomass or from synthesis gas (hydrogen, H2; CO2, carbon monoxide, CO). In anaerobes, common pathways involve the reduction of the intermediate acetyl-CoA with NAD(P)H by aldehyde (ALDH) and alcohol dehydrogenases (ADH). Alternatively, alcohols may be produced by the direct reduction of externally added or intermediately produced organic acids with reduced ferredoxin (Fdred). The key enzyme catalyzing this thermodynamically difficult reaction is aldehyde:ferredoxin oxidoreductase (AOR), an oxygen sensitive protein present in some anaerobic bacteria and archaea. Here, we present increasing evidence for the importance of the AOR-ADH pathway in alcohol producing anaerobes. AOR heavily depends on compounds with a low redox potential, and reactions potentially coupled to the pathway are discussed. The putative ancient AOR-ADH pathway may be relatively widespread among anaerobes, and it may play an important role in a sustainable bioenergy concept via the reduction of organic acids to their corresponding alcohols.

Reclaiming futures: communities helping teens overcome drugs, alcohol and crime--a new practice framework for juvenile justice.

Increasing numbers of youth entering the juvenile justice system on drug-related charges require new practice frameworks for youth advocates. Screening, assessment and intervention capabilities have not kept pace with the youth who need them. This article presents an overview of a new approach, based on the system of care movement, to integrated care service provision for this population. Utilizing the Reclaiming Futures initiative--being piloted in 10 communities in the United States--as a model for change, the article suggests that juvenile justice, in partnership with treatment and other community stakeholders, can have a positive impact on this social problem.

Moving mountains together: strategic community leadership and systems change.

Traditional leadership practice frameworks to guide systems change often fall short in today's practice environment. Reclaiming Futures is a national initiative to create an integrated, comprehensive, seamless system of care for teens with substance abuse problems involved in juvenile justice. It uses leadership and systems reform strategies to improve public health institutions. The premise is that interruption of the destructive cycle of drugs and delinquency can only be attained through the cultivation of shared strategic leadership. This article presents the limitations of traditional practice frameworks of leadership, describes the theory of this new approach to leadership development, and clarifies collaborative leadership and system change. The article includes an example of the framework in action, a discussion of lessons learned, and recommendations for community leadership development initiatives.

Systems of care for treatment of adolescent substance use disorders: background, principles and opportunities.

Adolescent substance abuse is a serious social problem facing the United States. Despite numerous recent advances in the clinical effectiveness of treatment approaches for this population, not enough attention has been paid to the adolescent treatment and service delivery infrastructures. The right services must be delivered through carefully organized and systematic community partnerships among agencies that serve the youth and families most in need. This article provides a working definition for the systems of care approach, reviews the movement's history within children's mental health services, addresses the feasibility of using the systems of care model for adolescent substance use disorders, and discusses principles and elements essential for successfully implementing a system of care for treatment of adolescent substance use disorders.