MC 3114

work:+1 212-854-2169

fax:+1 212-932-8273

kn5@columbia.edu

Research Summary

Our group deals with structural aspects of bioactive compounds and their mode of action. In most cases this involves investigating the interaction of small molecules with their biopolymeric receptors. We also continue to pursue isolation and characterization of natural products that are challenging because of their extremely limited quantity, instability, etc. Parallel with these studies, we place emphasis on developing new methods to facilitate structural studies, including biopolymers, as well as small molecules and new isolation methods.

A chemistry unit with two newly consstructed laboratories has recently started in Biosphere 2 in Arizona. The world in this huge glass-domed building with its rain forest, savannah, and ocean constitues an unprecedented "Nature" with no ultravioled light, and is bound to lead to new natural products chemistry.

1. Vision. (i) Retinal proteins. The changes occurring between the retinal chromophore (Rh) and rhodopsin (Rh, a G-protein coupled receptor) throughout the visual transduction was clarified in 2000. We are currently determining the Rh extramembrane loops and G protein interactions via photocross-linking. (ii) Affinity chromatography of the retinal pigment epithelium proteins using a biotin-linked retinoid has led to the isolation of three critical enzymes involved in the visual cycle. (iii) Ocular pigments are involved in age-related macular degeneration (AMD), for which no clear-cut remedy exists. After isolation from human eyes of fluorophores A2E and isoA2E consisting of two molecules of “vitamin A” and ethanolamine, we have identified two more fluorophores, a Rh-tris-A2E / Rh complex, and an all-trans retinal dimer. Intracellular as well as extracellular irradiation of these pigments with blue light leads to highly apoptotic oxiranes containing 9 additional oxygens. This singlet oxygen oxidation can be prevented by blue berry, the active constituent being the anthocyanines.
2. Ginkgolides. Extracts of the "fossil tree" Ginkgo biloba are reputed to improve memory and sharpen mental alertness. This bioactivity has been attributed to the cage molecules ginkgolides. They act as potent antagonists of the platelet-activating factor receptor (PAFR, a GPCR). Our aim is to study the neuromodulatory properties by determining the binding of ginkgolides to PAFR with cloned PAFR and in vivo and ex vivo studies of the central nervous system (CNS) by positron emission tomography (PET). Recently we have also found that gingkolides indeed prevent aggregation of the amyloid peptides that lead to Alzheimer’s disease.
3. Circular dichroism (CD) studies directed by Nina Berova (senior research scientist, research professor). Absolute configurational and conformational studies of biopolymers (DNA, Rh) by exciton-coupled CD, a nonempirical and versatile solution protocol. This extremely sensitive method can be performed at low microgram or even nanogram levels. New developments include absolute configurational assignments based on host-guest complexation of chiral(guests) to an auxiliary dimeric zinc porphyrin tweezer (host), and a micro-scale cross-metathesis reaction for introducing new chromphores. Theoretical studies have led to clarification of the porphyrin/porphyrin interactions and molecular recognition process. Ongoing studies on newly developed instrumentation for FDCD shows that fluorescence detected CD free of photoselection provides a valuable selectivity and several orders of magnitudes higher sensitivity of detection than conventional CD. Recent stereochemical studies include ab initio calculation of CD and ORD (optical rotatory dispersion) of natural products.
4. Development of mass spectrometric protocols to facilitate photoaffinity labeling and sequencing of sticky membrane proteins, using a wide variety of MS techniques and instrumentations, directed by Yasuhiro Itagaki (head, MS core facilities).

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Isolation and Characterization of a Retinal Pigment Epithelial Cell Fluorophores: An All-trans-retinal Dimer Conjugate, N.E. Fishkin, J.R. Sparrow, R. Allikmets, K. Nakanishi, Proc. Natl. Acad. Sci., 102, 7091-7096 (2005)

Anthocyanins protect against A2E Photooxidation and Membrane Permeabilization in Retinal Pigment Epithelian Cells, Y. P. Jang , J. Zhou, K. Nakanishi, J.R. Sparrow, Photochem. Photobiology, 81, 529-536 (2005)

A Cationic Zinc Porphyrin as a Chiroptical Probe for Z-DNA, M. Balaz, M. De Napoli, A. E. Holmes, A. Mammana, K. Nakanishi, N. Berova , R. Purrello, Angew. Chem. Int. Ed., 41, 4006-4009 (2005)

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