title: The importance of surface loops for stabilizing an eightfold beta alpha barrel protein.
abstract: An important step in understanding how a protein folds is to determine those regions of the sequence that are critical to both its stability and its folding pathway. We chose phosphoribosyl anthranilate isomerase from Escherichia coli, which is a monomeric representative of the (beta alpha)8 barrel family of proteins, to construct a variant that carries an internal tandem duplication of the fifth beta alpha module. This (beta alpha)9 variant was enzymically active and therefore must have a wild-type (beta alpha)8 core. It had a choice a priori to fold to three different folding frames, which are distinguished by carrying the duplicated segment as an insert into one out of three different loops. Steady-state kinetic constants, the fluorescence properties of a crucial tryptophan residue, and limited proteolysis showed that the stable (beta alpha)9 variant carries the insertion between beta-strand 5 and alpha-helix 5. This preference can be explained by the important role of loops between alpha helices and beta strands in stabilizing the structure of the enzyme.
authors.affiliation: Abteilung Biophysikalische Chemie, Universität Basel, Switzerland.
authors.forename: R
authors.initials: R
authors.lastname: Urfer
authors.forename: K
authors.initials: K
authors.lastname: Kirschner
mesh_terms: D000595:Amino Acid Sequence; D000877:Anthranilate Phosphoribosyltransferase / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology*; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D017510:Protein Folding; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D012091:Repetitive Sequences, Nucleic Acid / Q000235:genetics; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet
chemical_list: D011994:Recombinant Proteins; D000877:Anthranilate Phosphoribosyltransferase