Kinetics of Ca Dissociation from Cod Parvalbumin Studied by Fluorescent Stopped-flow Method.

Background: Small Ca-binding protein parvalbumin possesses two strong Ca/Mg- binding sites located within two EF-hand domains. Most parvalbumins have no tryptophan residues, while cod protein contains a single tryptophan residue, which fluorescence (spectrum maximum position and fluorescence quantum yield) is highly sensitive to the Ca association/dissociation.

Objective: Intrinsic protein fluorescence of cod parvalbumin can be used for elucidating the mechanism of Ca binding to this protein.

The Highly Conservative Cysteine of Oncomodulin as a Feasible Redox Sensor.

Oncomodulin (Ocm), or parvalbumin β, is an 11-12 kDa Ca-binding protein found inside and outside of vertebrate cells, which regulates numerous processes via poorly understood mechanisms. Ocm consists of two active Ca-specific domains of the EF-hand type ("helix-loop-helix" motif), covered by an EF-hand domain with inactive EF-hand loop, which contains a highly conservative cysteine with unknown function. In this study, we have explored peculiarities of the microenvironment of the conservative Cys18 of recombinant rat Ocm (rWT Ocm), redox properties of this residue, and structural/functional sensitivity of rWT Ocm to the homologous C18S substitution.

Comprehensive analysis of the roles of 'black' and 'gray' clusters in structure and function of rat β-parvalbumin.

Recently we found two highly conserved structural motifs in the proteins of the EF-hand calcium binding protein family. These motifs provide a supporting scaffold for the Ca binding loops and contribute to the hydrophobic core of the EF-hand domain. Each structural motif forms a cluster of three amino acids called cluster I ('black' cluster) and cluster II ('grey' cluster).

The impact of alpha-N-acetylation on structural and functional status of parvalbumin.

The effect of alpha-N-acetylation (Nt-acetylation) on the properties of parvalbumin (PA), a Ca2+-binding relaxing factor of skeletal muscles and major food allergen, has been explored. Intact PA contains an N-terminal acetyl group which is absent in the protein expressed in Escherichia coli (rWT), as confirmed by mass spectrometry. Compared to intact pike α-PA, its rWT form exhibits essentially altered profile of thermal unfolding, lowered α-helicity, and decreased affinities to Ca2+ and Mg2+.

Folding kinetics and structure of OEP16.

The chloroplast outer membrane contains different, specialized pores that are involved in highly specific traffic processes from the cytosol into the chloroplast and vice versa. One representative member of these channels is the outer envelope protein 16 (OEP16), a cation-selective high conductance channel with high selectivity for amino acids. Here we study the mechanism and kinetics of the folding of this membrane protein by fluorescence and circular dichroism spectroscopy, using deletion mutants of the two single tryptophanes Trp-77-->Phe-77 and Trp-100-->Phe-100.