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Wiley, European Journal of Biochemistry, 21(268), p. 5541-5549, 2001

DOI: 10.1046/j.1432-1033.2001.02491.x

Wiley, European Journal of Biochemistry, 21(268), p. 5541-5549

DOI: 10.1046/j.0014-2956.2001.02491.x

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Thermodynamic and kinetic analysis of porphyrin binding to Trichosantes cucumerina seed lectin

Journal article published in 2001 by Roopa Kenoth, D. Raghunath Reddy, Bhaskar G. Maiya, Musti J. Swamy
This paper is made freely available by the publisher.
This paper is made freely available by the publisher.

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Abstract

The interaction of several metallo-porphyrins with the galactose-specific lectin from Trichosanthes cucumeirna (TCSL) has been investigated. Difference absorption spectroscopy revealed that significant changes occur in the Soret band region of the porphyrins upon binding to TCSL and these changes have been monitored to obtain association constants (Ka) and stoichiometry of binding (n). The dimeric lectin binds two porphyrin molecules and the presence of the specific saccharide lactose did not affect porphyrin binding significantly, indicating that the sugar and the porphyrin bind at different sites. The Ka values obtained for the binding of different porphyrins with TCSL at 25 degrees C were in the range of 2 x 10(3)-5 x 10(5) m(-1). Association constants for meso-tetra(4-sulphonatophenyl)porphyrinato copper(II) (CuTPPS), a porphyrin bearing four negative charges and meso-tetra(4-methylpyridinium)porphyrinato copper(II) (CuTMPyP), a porphyrin with four positive charges, were determined at several temperatures; from the temperature dependence of the association constants, the thermodynamic parameters change in enthalpy (DeltaH degrees ) and change in entropy (DeltaS degrees ) associated with the binding process were estimated. The thermodynamic data indicate that porphyrin binding to TCSL is driven largely by a favourable entropic contribution; the enthalpic contribution is very small, suggesting that the binding process is governed primarily by hydrophobic forces. Stopped-flow spectroscopic measurements show that binding of CuTMPyP to TCSL takes place by a single-step process and at 20 degrees C, the association and dissociation rate constants were 1.89 x 10(4) m(-1).s(-1) and 0.29 s(-1), respectively.