Fluorimetric probes of the individual and competitive binding of 1-anilinonaphthalene-8-sulfonate, eosine and fluorescene to bovine serum albumin.

Fluorescence of 1-anilinonaphthalene-8-sulfonate (ANS) is greatly enhanced on its binding to bovine serum albumin (BSA). Fluorimetric titration shows that three ANS molecules bind per BSA molecule. The enhanced fluorescence of BSA-ANS is quenched by eosine (EOS); and one EOS physically displaces one ANS bound to BSA. The enhanced fluorescence of free ANS in the hydrophobic environment of the nonionic surfactant Triton X 100 is also quenched by EOS but by an energy transfer mechanism. The dye fluorescene (FLSN) also quenches the fluorescence of BSA-bound ANS, but by the energy transfer mechanism. The binding region of ANS in BSA has been speculated.

Spectrophotometric study of the relative affinities of teichoic acid for different metal ions.

Spectrophotometric study of the relative affinities of teichoic acid (TA) for Na+, Ca2+, Mg2+, Cd2+, Hg2+, Cu2+ and Pb2+ described on the basis of the disruption of metachromasia of dye 1,9-dimethyl methylene blue (DMMB)-polyanion (TA) system has revealed the following sequence of relative affinities of TA for these metals: Na+ < Ca2+ < or = Mg2+ < Cd2+ < Hg2+ < Cu2+ < Pb2+. Some of the metal ions particularly Pb2+ and Hg2+ have specific interactions with the dye, and induce distinct metachromatic band of the dye even in the absence of the polyanion.

Spectroscopic probe of distribution of the non-intercalating drug netropsin between DNA and heparin.

Circular dichroism has been used as a monitoring tool to probe the distribution of the non-intercalating drug netropsin (NTPS) between the two biomolecules DNA and heparin. The stoichiometry of the interaction of the individual biomolecules and the drug is determined from conductometric titrations; the titration in each case shows two breaks corresponding to two stoichiometries of interaction. Though netropsin is non-intercalating, DNA wins over heparin in binding the drug due to strong hydrogen bonding capability of NTPS in the minor grooves of DNA through its greater than NH donor groups. Potential hydrogen bond breakers like KF and urea reduce the induced dichroism of NTPS-DNA system, probably dislodging some drug from DNA through hydrogen bond breaking.

Induction of metachromasia and circular dichroism in the dye 1,9-dimethyl methylene blue by S. aureus wall teichoic acid.

Teichoic acid (TA) isolated from the Gram-positive bacteria Staphylococcus aureus induces blue shifted metachromasia in the dye 1,9-dimethyl methyleneblue (DMMB). Results of conductometric titrations show that DMMB binds to TA with 1:1 stoichiometry. Unlike DNA, this TA does not induce a definite metachromatic band in the dye acridine orange. S. aureus TA also induces strong circular dichroism (CD) in this dye with molar ellipticity value of the order of 10(4). This induction of CD in DMMB supports our earlier inference that TA in dilute solution at neutral pH has a helical conformation, and is not a random coil polymer.

Induction of metachromasia and circular dichroism in the dye 1,9-dimethyl methylene blue by ATP.

The theoretical prediction of induction of metachromasia [V Czikkely, H D Foersterling & H Kuhn (1970), Chem Phys Lett, 6,207] in a dye by a polyanion having only four to six anionic sites is proved experimentally, for the first time, in ATP--1.9-dimethyl methylene blue system. The findings show that ATP induces metachromasia in the dye at neutral pH, when ATP molecule remains fully charged providing four anionic sites to the dye cations. Conductometric titration shows that the dye molecules bind stoichiometrically to ATP (four dyes/ATP). However ATP at acidic pH and ADP and AMP at any pH fail to induce metachromasia. This is also the first report of induction of circular dichroism in bound dyes by ATP. Though the chiral moiety of ribose sugar in ATP may induce dichroism in the bound achiral dyes, the observed high molar ellipticity values indicate aggregation of bound dyes with twist in one sense initiated by the twisted conformation of the triphosphate chain in ATP. This inference on the state of conformation of ATP in its native environment is in agreement with that derived from PMR and spin lattice relaxation technique. It is thus interesting that the conformation of crystalline disodium ATP, as concluded from X-ray crystallography, is maintained by tetrasodium ATP in dilute aqueous solution--the native environment of ATP.

Spectrophotometric and spectrofluorometric titrations of teichoic acid.

Spectrophotometric and spectrofluorometric titrations of two strongly aggregating dyes, 1,9-dimethylmethylene blue (DMMB) and acridine orange (AO), by three anionic biopolymers, chondroitin sulphate A, DNA and teichoic acid (TA), have been described. Though the three polymers differ in their dye binding efficiencies and TA is a weak chromotrope, the equivalent weights of the polymers can be estimated accurately by these methods using the two dyes. Results show DMMB to be the preferred dye for spectrophotometric titration. The titrations can be used to estimate the equivalent weights of anionic polymers, and also for the quantitative estimation of such polymers of known equivalent weights.