title: Native-like in vivo folding of a circularly permuted jellyroll protein shown by crystal structure analysis.
abstract: A jellyroll beta-sandwich protein, the Bacillus beta-glucanase H(A16-M), is used to probe the role of N-terminal peptide regions in protein folding in vivo. A gene encoding H(A16-M) is rearranged to place residues 1-58 of the protein behind a signal peptide and residues 59-214. The rearranged gene is expressed in Escherichia coli. The resultant circularly permuted protein, cpA16M-59, is secreted into the periplasm, correctly processed, and folded into a stable and active enzyme. Crystal structure analysis at 2.0-A resolution, R = 15.3%, shows cpA16M-59 to have a three-dimensional structure nearly identical with that of the parent beta-glucanase. An analogous experiment based on the wild-type Bacillus macerans beta-glucanase, giving rise to the circularly permuted variant cpMAC-57, yields the same results. Folding of these proteins, therefore, is not a vectorial process depending on the conformation adopted by their native N-terminal oligopeptides after ribosomal synthesis and translocation through the cytoplasmic membrane.
authors.affiliation: Max-Delbrück-Centrum für Molekulare Medizin, Berlin, Germany.
authors.forename: M
authors.initials: M
authors.lastname: Hahn
authors.forename: K
authors.initials: K
authors.lastname: Piotukh
authors.forename: R
authors.initials: R
authors.lastname: Borriss
authors.forename: U
authors.initials: U
authors.lastname: Heinemann
mesh_terms: D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology; D001483:Base Sequence; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray / Q000379:methods; D017931:DNA Primers; D004795:Enzyme Stability; D015321:Gene Rearrangement; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010957:Plasmids; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D017931:DNA Primers; D011994:Recombinant Proteins; D006026:Glycoside Hydrolases; C028020:licheninase