title: Centrin: its secondary structure in the presence and absence of cations.
abstract: Centrin is a low molecular mass (20 kDa) protein that belongs to the EF-hand superfamily of calcium-binding proteins. Local and overall changes were investigated for interactions between cations and Chlamydomonas centrin using Fourier transform infrared (FT-IR) and circular dichroic (CD) spectroscopies. FT-IR spectral features studied included the amide I' band and the side-chain absorbances for aspartate residues located almost exclusively at the calcium-binding sites in the spectral region of 1700-1500 cm(-1). The amide I' band is exquisitely sensitive to changes in protein secondary structure and is observed to shift from 1626.5 to 1642.7 cm(-1) in the presence and absence of calcium. These spectral bands are complex and were further studied using two-dimensional Fourier transform infrared (2D-FT-IR) correlation along with curve-fitting routines. Using these methods the secondary structure contributions were determined for holocentrin and apocentrin. The alpha-helical content in centrin was determined to be 60%-53% in the presence and absence of cations, respectively. Furthermore, the beta-strand content was determined to be 12%-36%, while the random coil component remained almost constant at 7%-13.5% in the presence and absence of cations, respectively. Changes in the side-chain band are mostly due to the monodentate coordination of aspartate to the cation. A shift of approximately 4 cm(-1) (for the COO- antisymmetric stretch in Asp) from 1565 to 1569 cm(-1) is observed for apocentrin and holocentrin, respectively. Thermal dependence revealed reversible conformational transition temperatures for apocentrin at 37 degrees C and holocentrin at 45 degrees C, suggesting greater stability for holocentrin.
authors.affiliation: Department of Chemistry, University of Puerto Rico Mayagüez Campus, Mayagüez, Puerto Rico 00681-9019, USA. be_pastrana@rumac.uprm.edu
authors.forename: Belinda
authors.initials: B
authors.lastname: Pastrana-Rios
authors.forename: Wanda
authors.initials: W
authors.lastname: Ocaña
authors.forename: Michelle
authors.initials: M
authors.lastname: Rios
authors.forename: German Lorenzo
authors.initials: GL
authors.lastname: Vargas
authors.forename: Ginny
authors.initials: G
authors.lastname: Ysa
authors.forename: Gregory
authors.initials: G
authors.lastname: Poynter
authors.forename: Javier
authors.initials: J
authors.lastname: Tapia
authors.forename: Jeffrey L
authors.initials: JL
authors.lastname: Salisbury
mesh_terms: D000818:Animals; D001059:Apoproteins / Q000737:chemistry; D001224:Aspartic Acid / Q000737:chemistry*; D002118:Calcium / Q000737:chemistry*; D002135:Calcium-Binding Proteins / Q000737:chemistry*; D002412:Cations / Q000737:chemistry; D002696:Chlamydomonas / Q000737:chemistry*; D002868:Chromosomal Proteins, Non-Histone / Q000737:chemistry*; D002942:Circular Dichroism; D011489:Protein Denaturation; D017433:Protein Structure, Secondary; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods
chemical_list: D001059:Apoproteins; D002135:Calcium-Binding Proteins; D002412:Cations; D002868:Chromosomal Proteins, Non-Histone; C056608:caltractin; D001224:Aspartic Acid; D002118:Calcium