Examining Fmoc‐amino acid derivatives as selective inhibitors of butyrylcholinesterase

Abstract

The design of enzyme inhibitors to act as drugs or as substrates for biocatalysts remains a challenge in biochemistry, pharmacology, and biotechnology. To mitigate the effects of Alzheimer's disease (AD), a neurodegenerative disease characterized by progressive memory loss and the sixth leading cause of death in the United States, numerous enzyme inhibitors have been developed. Activity of the enzyme butyrylcholinesterase (BChE) increases in Alzheimer's patients, and this increase is suggested to lead to a depletion of the neurotransmitter, acetylcholine. To counter this increase in activity BChE‐specific inhibitors have been developed to manage the progression of AD. We hypothesized 9‐fluorenylmethyloxycarbonyl (Fmoc) amino acids could provide a new class of inhibitors, as these compounds features found in BChE inhibitors currently available as pharmaceuticals. The readily available chemistry makes amino acids an attractive scaffold for identifying BChE inhibitors. We found Fmoc‐Leu‐O− as an effective and selective BChE inhibitor with an inhibition constant (KI) value of 115 μM. A series of Fmoc‐Leu‐O− derivatives with side chain modifications were then evaluated. The Fmoc‐amino acids were purchased or synthesized from the corresponding amino acid. The KI value for each compound was determined using kinetics assays and UV‐absorbance spectroscopy. For the four Fmoc‐Leu‐O− derivatives tested, Fmoc‐neopentylglycine‐O− showed the greatest potency with an inhibition constant of 36 μM, ~three‐fold lower than the original compounds. A series of Fmocdi‐, tri‐, tetra‐, and pentapeptides were synthesized by solid phase peptide synthesis. The products were purified by HPLC and characterized by NMR and mass spectrometry. Our initial results identified the dipeptide (Fmoc‐Leu‐Leu‐O−) as an effective inhibitor with KI value of 28 μM. We are currently investigating if longer Fmoc‐Leu‐O− peptides and other Fmoc‐peptides are potent and selective BChE inhibitors. The results explore a new class of potent and selective BChE inhibitors and may identify features important for binding in the enzyme active site.Support or Funding InformationThis project was supported by the National Institute of General Medical Sciences, National Institutes of Health (NIH) under award numbers T34GM008074, UL1GM118979, and RL5GM118978, and the National Science Foundation (NSF; MRI CHE‐1337559). The content is solely the responsibility of the authors and does not necessarily represent the official views of the NIH or NSF.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.