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The cation-? interaction between Lys53 and the flavin of fructosamine oxidase (FAOX-II) is critical for activity.

ABSTRACT: Fructosamine oxidases (FAOXs) are flavin-containing enzymes that catalyze the oxidative deglycation of low molecular weight fructosamines or Amadori products. The fructosamine substrate is oxidized by the flavin in the reductive half-reaction, and the reduced flavin is then oxidized by molecular oxygen in the oxidative half-reaction. The crystal structure of FAOX-II from Aspergillus fumigatus reveals a unique interaction between Lys53 and the isoalloxazine. The ammonium nitrogen of the lysine is in contact with and nearly centered over the aromatic ring of the flavin on the si-face. Here, we investigate the importance of this unique interaction on the reactions catalyzed by FAOX by studying both half-reactions of the wild-type and Lys53 mutant enzymes. The positive charge of Lys53 is critical for flavin reduction but plays very little role in the reaction with molecular oxygen. The conservative mutation of Lys53 to arginine had minor effects on catalysis. However, removing the charge by replacing Lys53 with methionine caused more than a million-fold decrease in flavin reduction, while only slowing the oxygen reaction by ?30-fold.

SUBMITTER: Collard F

PROVIDER: S-EPMC3557951 | biostudies-literature | 2011 Sep

REPOSITORIES: biostudies-literature

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The cation-π interaction between Lys53 and the flavin of fructosamine oxidase (FAOX-II) is critical for activity.

Collard François F Fagan Rebecca L RL Zhang Jianye J Nemet Ina I Palfey Bruce A BA Monnier Vincent M VM

Biochemistry 20110825 37

Fructosamine oxidases (FAOXs) are flavin-containing enzymes that catalyze the oxidative deglycation of low molecular weight fructosamines or Amadori products. The fructosamine substrate is oxidized by the flavin in the reductive half-reaction, and the reduced flavin is then oxidized by molecular oxygen in the oxidative half-reaction. The crystal structure of FAOX-II from Aspergillus fumigatus reveals a unique interaction between Lys53 and the isoalloxazine. The ammonium nitrogen of the lysine is ...[more]

PMID: 21755947

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Project description:The chemistry community now recognizes the cation-π interaction as a major force for molecular recognition, joining the hydrophobic effect, the hydrogen bond, and the ion pair in determining macromolecular structure and drug-receptor interactions. This Account provides the author's perspective on the intellectual origins and fundamental nature of the cation-π interaction. Early studies on cyclophanes established that water-soluble, cationic molecules would forego aqueous solvation to enter a hydrophobic cavity if that cavity was lined with π systems. Important gas phase studies established the fundamental nature of the cation-π interaction. The strength of the cation-π interaction (Li(+) binds to benzene with 38 kcal/mol of binding energy; NH4(+) with 19 kcal/mol) distinguishes it from the weaker polar-π interactions observed in the benzene dimer or water-benzene complexes. In addition to the substantial intrinsic strength of the cation-π interaction in gas phase studies, the cation-π interaction remains energetically significant in aqueous media and under biological conditions. Many studies have shown that cation-π interactions can enhance binding energies by 2-5 kcal/mol, making them competitive with hydrogen bonds and ion pairs in drug-receptor and protein-protein interactions. As with other noncovalent interactions involving aromatic systems, the cation-π interaction includes a substantial electrostatic component. The six (four) C(δ-)-H(δ+) bond dipoles of a molecule like benzene (ethylene) combine to produce a region of negative electrostatic potential on the face of the π system. Simple electrostatics facilitate a natural attraction of cations to the surface. The trend for (gas phase) binding energies is Li(+) > Na(+) > K(+) > Rb(+): as the ion gets larger the charge is dispersed over a larger sphere and binding interactions weaken, a classical electrostatic effect. On other hand, polarizability does not define these interactions. Cyclohexane is more polarizable than benzene but a decidedly poorer cation binder. Many studies have documented cation-π interactions in protein structures, where lysine or arginine side chains interact with phenylalanine, tyrosine, or tryptophan. In addition, countless studies have established the importance of the cation-π interaction in a range of biological processes. Our work has focused on molecular neurobiology, and we have shown that neurotransmitters generally use a cation-π interaction to bind to their receptors. We have also shown that many drug-receptor interactions involve cation-π interactions. A cation-π interaction plays a critical role in the binding of nicotine to ACh receptors in the brain, an especially significant case. Other researchers have established important cation-π interactions in the recognition of the "histone code," in terpene biosynthesis, in chemical catalysis, and in many other systems.

Project description:Nicotine addiction begins with high-affinity binding of nicotine to acetylcholine (ACh) receptors in the brain. The end result is over 4,000,000 smoking-related deaths annually worldwide and the largest source of preventable mortality in developed countries. Stress reduction, pleasure, improved cognition and other central nervous system effects are strongly associated with smoking. However, if nicotine activated ACh receptors found in muscle as potently as it does brain ACh receptors, smoking would cause intolerable and perhaps fatal muscle contractions. Despite extensive pharmacological, functional and structural studies of ACh receptors, the basis for the differential action of nicotine on brain compared with muscle ACh receptors has not been determined. Here we show that at the alpha4beta2 brain receptors thought to underlie nicotine addiction, the high affinity for nicotine is the result of a strong cation-pi interaction to a specific aromatic amino acid of the receptor, TrpB. In contrast, the low affinity for nicotine at the muscle-type ACh receptor is largely due to the fact that this key interaction is absent, even though the immediate binding site residues, including the key amino acid TrpB, are identical in the brain and muscle receptors. At the same time a hydrogen bond from nicotine to the backbone carbonyl of TrpB is enhanced in the neuronal receptor relative to the muscle type. A point mutation near TrpB that differentiates alpha4beta2 and muscle-type receptors seems to influence the shape of the binding site, allowing nicotine to interact more strongly with TrpB in the neuronal receptor. ACh receptors are established therapeutic targets for Alzheimer's disease, schizophrenia, Parkinson's disease, smoking cessation, pain, attention-deficit hyperactivity disorder, epilepsy, autism and depression. Along with solving a chemical mystery in nicotine addiction, our results provide guidance for efforts to develop drugs that target specific types of nicotinic receptors.

Dataset Information

The cation-? interaction between Lys53 and the flavin of fructosamine oxidase (FAOX-II) is critical for activity.

Publications

The cation-π interaction between Lys53 and the flavin of fructosamine oxidase (FAOX-II) is critical for activity.

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