B ring regulation of colchicine binding kinetics and fluorescence.

Bhattacharyya, Bhabatarak; Howard, R.; Maity, Sankar N.; Brossi, Arnold; Sharma, Padam N.; Wolff, J.

doi:10.1073/pnas.83.7.2052

Cited by 60 publications

(45 citation statements)

References 27 publications

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“…Previously, it was observed that this time dependence is related to the presence of substituents in the B ring, especially the C=O group at C7 of the colchicine nucleus [8]. Colcemid, a colchicine analogue which lacks the C=O group, binds tubulin much faster than colchicine [13].…”

Section: Time Dependence Of Nbd-colcemid Bindingmentioning

confidence: 99%

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N‐(7‐Nitrobenz‐2‐oxa‐1,3‐diazol‐4‐yI)colcemid, a probe for different classes of colchincine‐binding site on tubulin

Sengupta

Puri

Surolia

et al. 1993

European Journal of Biochemistry

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The nature of binding of 7-nitrobenz-2-oxa-l,3-diazol-4-yl-colcemid (NBD-colcemid), an environment-sensitive fluorescent analogue of colchicine, to tubulin was tested. This article reports the first fluorometric study where two types of binding site of a colchicine analogue on tubulin were detected. Binding of NBD-colcemid to one of these sites equilibrates slowly. NBD-colcemid competes with colchicine for this site. Binding of NBD-colcemid to this site also causes inhibition of tubulin self-assembly. In contrast, NBD-colcemid binding to the other site is characterised by rapid equilibration and lack of competition with colchicine. Nevertheless, binding to this site is highly specific for the cholchicine nucleus, as alkyl-NBD analogues have no significant binding activity. Fast-reaction-kinetic studies gave 1.76X105 M-' s-' for the association and 0.79 s-l for the dissociation rate constants for the binding of NBD-colcemid to the fast site of tubulin. The association rate constants for the two phases of the slow site are 0.016X10-4 M-' s-' and 3.5X

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Section: Time Dependence Of Nbd-colcemid Bindingmentioning

confidence: 99%

“…The association is a slow, poorly reversible process which is highly temperature dependent [5-71. It has been argued that rings B and C of the colchicine molecule are responsible for such unusual properties [8]. The kinetics of colchicine/tubulin interaction were found to consist of two steps, as described by the following scheme [9]: T+C*[TC]+TC*.…”

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confidence: 99%

N‐(7‐Nitrobenz‐2‐oxa‐1,3‐diazol‐4‐yI)colcemid, a probe for different classes of colchincine‐binding site on tubulin

Sengupta

Puri

Surolia

et al. 1993

European Journal of Biochemistry

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“…However, the first-order dissociation rates for DAACtubulin and aminocolchicinoid-tubulin complexes are comparable and are about 100 -150-fold higher than that of the colchicine-tubulin complex. It was suggested previously that the ϾCϭO in the side chain is responsible for the poor reversibility of the colchicine-tubulin interaction (19). Dissociation rate of another colchicine analog wherein ϾCϭO was substituted by ϾCϭS (colchicine fluorescein) (see Fig.…”

Section: Resultsmentioning

confidence: 99%

Thermodynamics of Colchicinoid-Tubulin Interactions

Chakrabarti

Sengupta

Bhattacharyya

1996

Journal of Biological Chemistry

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The quenching of tryptophan fluorescence has been used to determine the kinetic and thermodynamic parameters of binding of B-ring analogs of colchicine to tubulin. The on rate, activation energy, off-rate, and thermodynamics of binding reaction have been found to be controlled at different points of analog structure. The on-rate and off-rate of deacetamidocolchicine (DAAC) binding with tubulin is 17 times slower than that of 2-methoxy-5-(2,3,4-trimethoxyphenyl)tropone-tubulin (AC-tubulin) interaction, although both reactions have very similar activation energies. The presence of B-ring alone does not significantly affect the thermodynamics of the binding reactions either, since both AC-tubulin and DAAC-tubulin interactions are enthalpy driven. Introduction of a NH 2 group at C-7 position of the B-ring, as in deacetylcolchicine (NH 2 -DAAC) lowers the on-rate further with a significant rise in the value of the activation energy. However, bulkier substitutions at the same position, as in demecolcine (NHMe-DAAC) and N-methyldemecolcine (NMe 2 -DAAC) have no significant additional effect either on the on-rate or on the value of activation energy. Introduction of NH 2 group in the C-7 position of B-ring also increases the positive entropy of the binding reaction to a significant extent, and it is maximum when NMe 2 is substituted instead of NH 2 group. Thus, interaction of NH 2 -DAAC, NHMe-DAAC, and NMe 2 -DAAC with tubulin are entropy driven. Our results suggest that the B-ring side chain of aminocolchicinoids makes contact(s) with dimeric tubulin molecules.Colchicine, the major alkaloid in Colchicum autumnale, is medically used for the treatment of gout (1) and Familial Mediterranean fever (2). Due to its immense therapeutic importance, a large number of colchicine and thiocolchicine analogs have been synthesized and tested for their biological activities (3-5). Colchicine exerts its antimitotic property upon binding to a high affinity site on the tubulin heterodimer (6 -8). It is composed of a trimethoxybenzene ring (A-ring), a methoxytropone ring (C-ring), and a seven-membered ring (B-ring), which anchors the A-and C-rings (Fig. 1). Structure activity studies indicate that the A-and C-rings of colchicine comprise the minimum structural features of the molecule necessary for its high affinity binding to tubulin. Insertion of a bulky group in the A-ring of colchicine, as in colchicoside, causes complete loss of binding, indicating that the requirement of the A-ring is stringent (4). On the other hand, several changes in the C-ring such as different substitutions at the C-10 position or a replacement of the seven-membered ring with a six-membered ring are tolerated (9 -14). Colchicine analogs modified at, or depleted of, the B-ring are known to retain potent antimitotic activity, self-assembly inhibitory activity, and the binding activity to tubulin at the colchicine site (15-17). Nevertheless, the presence of B-ring alone, or substituents at C-7 position, influences the on-rate, activation energy, off-rate, reversi...

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“…The importance of the B-ring of colchicine in its binding to tubulin has been emphasized by different laboratories [1][2][3][4][5][6][7][8]. It has been observed that several properties of the colchicine-tubulin interaction such as the association rate, reversibility and the promotion of drug fluorescence are related to the B-ring of colchicine [7].…”

Section: Introductionmentioning

confidence: 99%

“…It has been observed that several properties of the colchicine-tubulin interaction such as the association rate, reversibility and the promotion of drug fluorescence are related to the B-ring of colchicine [7]. Recently significant differences in the binding and thermodynamic parameters such as activation energy, entropy and enthalpy have been reported for colehicine and 2-methoxy-5-(2' ,3' ,4' -trimethoxyphenyl)tropone (called AC because it lacks the B-ring of colchicine, see fig.l) [5,6].…”

Section: Introductionmentioning

confidence: 99%

Properties of B‐ring analogues of colchicine

Maity

Bhattacharyya

1987

FEBS Letters

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Absorption spectra of colchicine and its analogues are affected by the presence of the B‐ring, although it is not part of the chromophore (C‐ring). Thus, 2‐methoxy‐5‐(2′,3′4′‐trimethoxyphenyl)tropone has absorption maxima at 341 nm, whereas that of desacetamidocolchicine is at 353 nm. A similar red shift in the λmax of colchicine, desacetamidocolchicine and 2‐methoxy‐(2′,3′,4′‐trimethoxyphenyl)tropone also occurs when they are immobilized in the binding site to tubulin or in pure glycerol. We also observed that the B‐ring of colchicine alone or with substituent does not affect the UV‐induced rearrangement of colchicine to lumicolchicine. However, in the absence of the B‐ring, as in the case of 2‐methoxy‐5‐(2′,3′4′‐trimethoxyphenyl)tropone, the rearrangement reaction of the C‐ring slows down significantly.

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B ring regulation of colchicine binding kinetics and fluorescence.

Cited by 60 publications

References 27 publications

N‐(7‐Nitrobenz‐2‐oxa‐1,3‐diazol‐4‐yI)colcemid, a probe for different classes of colchincine‐binding site on tubulin

N‐(7‐Nitrobenz‐2‐oxa‐1,3‐diazol‐4‐yI)colcemid, a probe for different classes of colchincine‐binding site on tubulin

Thermodynamics of Colchicinoid-Tubulin Interactions

Properties of B‐ring analogues of colchicine

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