Welcome to the Miller Laboratory Web Pages 

Group Photo, October 2017. R.Sarma photo.

Group Photo, October 2017. R.Sarma photo.

Group Photo, March 2017 (Absent TW and JR).

Group Photo, July 2015.

Anne-Frances Miller

Professor of Chemistry, and of Biochemistry Download CV.

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The Miller group studies  Enzymatic Redox Catalysis.



There is no planet B. We must adopt ways to live lighter on our precious earth. Fortunately many new technologies are available to help us consume less energy and water while also living higher-quality lives. However implementation remains a challenge. The Miller lab seeks to make solutions from nature practical for industrial and domestic use. Out twin foci are energy and water. In both cases, we turn to enzymes. We are learning about energy conservation at the level of electrons from enzymes that employ a recently-appreciated mechanism to enable organisms to flourish in meager environments. We are also putting enzymes to work, cleaning up used water so that it can be used again, and again, and again....Click here to learn more.  


1. Superoxide Dismutase Projects.

When the enzyme you have is not quite the enzyme you want, we turn to enzyme engineering. Good tools and understanding exit for altering substrate specificity, but modification of activity has proven much more difficult to accomplish without disrupting the enzyme. Non-heme iron (Fe) is used in a wide variety of enzymes to effect a stunning array of different reactions, but particularly reactions involving activation of O2. Therefore we have employed Fe-containig superoxide dismutase (SOD) and its close cousin the Mn-specific SOD of E. coli [72] to identify principles of redox tuning and manipulate the active site to produce variants with reduction midpoint potentials (Eo) ranging over 0.9 V (21 kCal/mol or 89 kJ/mol), without disrupting the protein [59]. Thus this system serves as a molecular laboratory for modifying a non-heme-Fe site to perform desired chemistry. Optical, EPR and NMR spectroscopy are used with Xray crystallography and titrations to obtain quantitative measures of fundamental elements of activity. Click here to learn more.  


2. Nitroreductase, a Promiscuous Enzyme that Loves Trash.

Nitroreductase is a promiscuous enzyme with promise for bioremediation of herbicides, pesticides, explosives and biproducts of textile industry. The enormous versatility of this flavo-enzyme suits it for many applications. We are also developing it for converting inexpensive nitroaromatic compounds into the corresponding aromatic hydroxylamines and amines, which are invaluable starting materials for synthesis of pharmaceuticals. We have explored the basis for NR's promiscuity, which breaks the 'rule' that each enzyme is selective for a given substrate. NMR reveals protein dynamics underlying NR's very broad substrate specificy range. Stopped-flow kinetics studies reveal an extremely simple mechanism depending primarily on the flavin and active site water molecules, and thus relatively independent of specific interactions with protein residues [82,88]. Click here to learn more.


3. Spectroscopic Elucidation of Flavin Control by Proteins.

When the enzyme we have is not quite the enzyme we want, we turn to enzyme engineering. Rational engineering of redox reactivity entails control of the electronics of a cofactor. Flavins are competent for an exceptionally broad range of chemistry, so flavoenzymes provide excellent potential for engineering. We are working to provide insight into how interactions in proteins modify the electronics of bound flavins. Indeed, chemically identical flavins can display contrasting reactivities when bound in different protein sites. 15N and 13C NMR chemical shift tensors are sensitive probes of the natures and relative energies of the frontier orbitals: the very orbitals that underlie the flavin's reactivity [55]. We have used NMR to learn how proteins modify the frontier orbitals of bound flavins via hydrogen bonds to N and O sites of the flavin [69]. Resonance Raman spectroscopy provides another very sensitive probe of the effects of individual hydrogen bonds and local charges, providing the possibility of understanding how the protein site produces altered flavin reactivity. Deployment of these methods in conjunction with mutagenesis is paving the way to intentionally altering the reactivity of a bound flavin via modification of its protein site. Click here to learn more


4. Bioremediation of Trash to Treasure.

In many parts of the world potable water has become sufficiently scarce that it is the object of political clashes and strife between neighbors. Even here, waste is accumulating in our watersheds causing disease and poisoning the ecosystems we rely upon. Many of the most troubling chemical wastes are present and toxic at very low concentrations making superfund-type approaches impractical. Enzymes offer a solution because they are active in water and able to bind substrates present at < mM concentrations even in complicated mixtures. We have demonstrated NR's capacity to transform veterinary antibiotics (waste streams of CAFOs) and capture energy as electrical current when the enzyme is immobilized on electrodes. Some wastes are sufficiently abundant in effluent streams that they represent a significant untapped oppurtunity to recapture energy and material [69]. We have demonstrated this using immobilized laccase to produce the valuable food additive vanillin from a compound that is a bi-product of the paper industry. Click here to learn more.


5. Contrasting Reactivities of the two Flavins in Bifurcating Electron Transfer Flavoproteins.

Electron transfer flavoproteins (Etfs) are heterodimeric flavoproteins found in every kingdom of life. They are 'plug-and-play' electron carriers that interface between diverse electron sources and the respiratory electron transfer chain (aerobes) or NAD(P)H and ferredoxins able to supply demanding reductive reactions (anaerobes). The former group of Etfs uses its single FAD to mediate 1-electron transfer and contains a 'structural' AMP whereas the latter group contains a second FAD proposed to undergo electron bifucation [93]. Flavin-based electron bifurcation is a recently-described third fundamental mechanism of energy conservation whereby the energy of two modestly-reducing electrons from NADH is redistributed among the two electrons to yield a more strongly reducing electron able to drive nitrogen fixation. Thus lessons learned in bifurcating Etf are applicable to a wide variety of reactions and the fundamental principles we seek to articulate can inform ongoing efforts to design novel energy efficient materials and devices.  Click here to learn more




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Update: Oct. 2017                                                                                               

Copyright 2017 A.-F. Miller     

Comments: A.-F. Miller