title: Sequential melting of two hydrophobic clusters within the green fluorescent protein GFP-cycle3.
abstract: The analysis of the three-dimensional structure of green fluorescent protein (GFP-cycle3) revealed the presence of two well-defined hydrophobic clusters located on the opposite sides of the GFP β-can that might contribute to the formation of partially folded intermediate(s) during GFP unfolding. The microcalorimetric analysis of the nonequilibrium melting of GFP-cycle3 and its two mutants, I14A and I161A, revealed that due to the sequential melting of the mentioned hydrophobic clusters, the temperature-induced denaturation of this protein most likely occurs in three stages. The first and second stages involve melting of a smaller hydrophobic cluster formed around the residue I161, whereas a larger hydrophobic cluster (formed around the residues I14) is melted only at the last GFP-cycle3 denaturation step or remains rather structured even in the denatured state.
authors.affiliation: Institute of Protein Research, Russian Academy of Sciences, 142290 Pushchino, Moscow Region, Russia.
authors.forename: Tatiana N
authors.initials: TN
authors.lastname: Melnik
authors.forename: Tatiana V
authors.initials: TV
authors.lastname: Povarnitsyna
authors.forename: Anatoly S
authors.initials: AS
authors.lastname: Glukhov
authors.forename: Vladimir N
authors.initials: VN
authors.lastname: Uversky
authors.forename: Bogdan S
authors.initials: BS
authors.lastname: Melnik
mesh_terms: D049452:Green Fluorescent Proteins / Q000737:chemistry*; D057927:Hydrophobic and Hydrophilic Interactions; D009154:Mutation; D011489:Protein Denaturation; D017510:Protein Folding; D013696:Temperature; D013816:Thermodynamics
chemical_list: D049452:Green Fluorescent Proteins