A Study of Structural Stability of Kinase-related
Protein (KRP)

T. L. Bushueva¹, M. V. Teplova², V. N. Bushuev¹, D. S. Kudryashov¹,
A. V. Vorotnikov¹, and V. P. Shirinskii¹

¹ Institute of Experimental Cardiology, Russian Cardiology Research Center, Moscow, Russia;
E-mail: bush@cardio.med.msu.su
² Bach Institute of Biochemistry, Russian Academy of Sciences, Moscow, 121552 Russia

Received April 14, 1998

Abstract—The structural stability of the smooth muscle kinase-related protein (KRP) was studied by various
methods: intrinsic (tryptophanyl) protein fluorescence, differential scanning calorimetry, and NMR. It was sug-
gested from the analysis of fluorescence and NMR spectra and fluorescence quenching that KRP is a compact
globular protein, with neither the carboxy- nor the aminoterminal high-mobility domains incorporated into the
globule. The only tryptophan residue of the molecule is inside the globule, inaccessible for water molecules and
poorly accessible for iodide and acrylamide. The lack of fluorescence quenching by Cs⁺ suggests that positively
charged groups are dominant in the vicinity of the tryptophan residue. Studies of the KRP molecule tolerance
to guanidine hydrochloride (GHC), pH, and temperature changes revealed that, in addition to native and
unfolded conformational states, there are intermediate conformational states, which are characterized by high
intramolecular mobility. Transition from native to an intermediate state occurs at moderately low concentrations
of GHC (<1.5 M), pH 8–10, and temperature close to physiological (<37°C). The intermediate conformational
structures are suggested to contribute to the functional activity of the protein.

Key words: kinase-related protein (KRP), protein conformation, structural stability, fluorescence, calorimetry,
NMR

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