Maurizio Zandomeneghi

Bovine lens aldose reductase: tight binding of the pyridine coenzyme

Analysis by HPLC of the protein-free supernatant obtained after denaturation of aldose reductase shows that the native form of the enzyme (ARb) contains a tightly bound NADP+, which is absent in the oxidatively modified form (ARa). The absorption, fluorescence, and circular dichroism spectra of ARb and ARa are consistent with the presence of the cofactor only in the native form of aldose reductase. On the other hand, the modified enzyme, in appropriate thiol reducing conditions, can tightly bind NADP+. This indicates a potential reversibility of the modification of aldose reductase, at least in terms of retention of the cofactor.

On the Chiroptical Behavior of Conjugated Multichromophoric Compounds of a New Pseudoaromatic Class: Bicolchicides and Biisocolchicides

Background It is well known that, stemming from the mutual interplay between chromophores, circular dichroism (CD) is a powerful technique to deal with structural problems for both the small organic molecule and the biopolymer. However, quantitative interpretations of the spectroscopic and structural terms that give rise to the exciton couplet are usually presented for ideal cases, or a few CD bands only are taken into account, overlooking the role of the solvent medium. Methodology/Principal Findings Circular dichroism and UV absorption spectra were carried out for colchicide (3) and isocolchicide (6), as well as their coupling products, 10,10′-bicolchicide (2) and 9,9′-biisocolchicide (5), in both hydrogen bonding and non hydrogen bonding solvents, as well as MeCN/H2O mixtures. A dramatic control by the solvent emerged, as even tiny changes in the composition of solvent mixtures, at ca 1 water molar fraction, induced a dramatic modification of their CD bands. A mutarotation phenomenon - long known for isocolchicine (8) - was also observed for 5, and can be attributed to the interconversion between atropisomers (R a,7S),(R a,7′S)-5a and (R a,7S),(S a,7′S)-5b. Conclusions/Significance Our data show that with molecules built on two structurally identical moieties which embody both hydrophilic and hydrophobic groups, even tiny changes in the composition of solvent mixtures cause a dramatic modification of the CD bands. Their analysis arrives at a qualitative rationalization of the observed CD couplets from the coupling of high energy transitions, while attempts at a quantitative interpretation of these phenomena through time-dependent density functional theory allowed to reproduce satisfactorily the CD spectrum in the 300–450 nm region only. Failure with higher energies probably reflects currently inadequate specific theoretical treatments of the solvent medium.

Isolation of atropisomers in both the isocolchicide and colchicide series of alkaloids and determination of their chiroptical properties

Abstract The isolation and dichroic behavior of atropisomeric colchicinoids is described here for the first time. The cases range from stable to rapidly interconverting atropisomeric couples.

Laser photochemistry: The intramolecular cyclization of carvone to carvonecamphor

Abstract Kr-ion laser irradiation of carvone ( 1 ) at 350.7 and 356.4 nm gave carvonecamphor ( 2 ) in a much higher yield (88%) than in conventional photochemistry, while ( 3 ) was a primary photo-by-product.

Determination of Holo- and Apo-Riboflavin Binding Protein in Avian Egg Whites through Circular Dichroism and Fluorescence Spectroscopy

The hen egg white contains proteins able to strongly bind, with a definite stoichiometry, small molecules such as biotin and riboflavin, or ions such as Cu2+ or Fe3+. The complexation process modifies the spectral properties of these low-molecular-weight species. On the basis of these changes, it is possible, in principle, to measure the quantity of the binding protein and to evaluate the protein-substrate interactions. Here, we present a method to determine the concentration of both the apo and holo forms of the riboflavin-binding protein (RFBP) present in avian egg white, by measuring the circular dichroism (CD) related to the controlled addition of riboflavin (or vitamin B2) to the egg white. At the same time, front-face fluorescence is used to confirm the concentration of apo-RFBP obtained from CD data. The method is based on data only from spectroscopy, and no process involving either extraction, chromatography, electrophoresis, or mass spectrometry is involved. We study the egg whites from four different avian species, reporting and comparing the concentration of the apo- and holo-RFBP and the molar circular dichroism spectra (Deltaepsilon) of riboflavin in the RFBP binding site. Finally, egg whites from different hen individuals are analyzed, and a surprising variation of the RFBP concentration is found.

Interaction of a reactive dye with serum albumins and with aminoacids: the dye as a chiral label

Abstract HSA and BSA, human and bovine serum albumins, react with a reactive dye, C.I. Reactive Blue 29 (levafix brillant blue EB) giving adducts containing two dye molecules per protein molecule. While cysteine was able to react with the dye, the cysteine(s) present in the proteins was not. The dye-protein adducts reveal strong circular dichroism in the 300–720 nm spectral region where only absorption bands of the achiral dye molecule are active. The intensity of these bands depends on the conformation of the protein: particularly the 530–720 nm band. On tuning the denaturation degree by means of SDS addition and removal, the intensity of this band varies between zero and a maximum when the protein is in fully denaturated and native conditions, respectively. This behavior evidences a chiral label function of the dye that can be useful in studying processes regarding proteins.

Changes in the photochemistry of tropone or 2-methoxytropone on complexation with acids

On addition of either BF3·Et2O or H2SO4 the valence photoisomerization of 2-methoxytropone (1) in CH3CN is diverted from 1-methoxy-(2) to 3-methoxybicyclo[3.2.0]hepta-3,6-dien-2-one (3) as products whereas with tropone there is a change from photodimerization to valence isomerization to give cleanly bicyclo[3.2.0]hepta-3,6-dien-2-one (5).

Chiroptical studies of labile or difficult to resolve molecules generated by chiral laser photochemistry.1. (1S,5R-1-methoxybicyclo[3.2.0] hepta-3,6-dien-2-one

Abstract The optical activity of (1 S ,5 R -1-methoxybicyclo[3.2.0]hepta-3,6-dien-2-one ( 1 ) is determined by partial photoresolution of the racemate with circularly polarized laser light; although a rearranged, optically active product was also formed, isolation of the 1 enantiomers was note required.

Photochemistry of some 2,5-substituted tropone mesogens

Abstract The photoreactivity of three 2-acyloxy-5-alkoxytropone derivatives, two of which are known to show thermotropic liquid crystalline phases, has been studied with the aim of finding photoresponsive mesogenic materials. The irradiation of solutions of these compounds in CH 3 CN under air atmosphere brings to endoperoxides which do not show mesomorphism and are not fully reversible to their respective precursors neither photochemically nor thermally. From the chemical point of view, the facile and relatively neat formation of endoperoxides open synthetic routes to new functionalized tropone derivatives.

Chiroptical studies of saturated ketones partially photoresolved by chiral laser photochemistry

Partial photoresolution of tricyclo[5.4.0.01,5]undecan-5-one ((±)-1) with circularly-polarised 333.6 nm light from an Ar-ion c.w. laser led to Δemax= -0.40 and Δe/emax = 1.8% at 300nm in n-hexane and 1S, 7R, 5R configuration for (-)-1.