title: Secondary structure characterization of beta-lactamase inclusion bodies.
abstract: The secondary structure of proteins in E. coli inclusion bodies was investigated via Raman spectroscopy. Inclusion bodies were purified from cells expressing different forms of RTEM beta-lactamase and grown at either 37 or 42 degrees C. All of the solid phase inclusion body samples examined gave amide I band spectra that were perturbed from that of the native, purified protein in both solution and powder forms; secondary structure estimates indicated significant decreases in alpha-helix and increases in beta-sheet contents in the inclusion body samples. The structure estimates for inclusion bodies isolated from 37 degrees C cultures were similar, regardless of aggregate localization in the E. coli cytoplasmic or periplasmic spaces or beta-lactamase precursor content. Inclusion bodies obtained from 42 degrees C cells exhibited a further reduction of alpha-helix and augmentation of beta-sheet contents relative to those from 37 degrees C cultures. These results are consistent with the paradigm for inclusion body formation via the self-association of intra-cellular folding intermediates having extensive secondary structure content. Further, the overall secondary structure content of inclusion bodies is not significantly affected by subcellular compartmentalization, but may be altered at increased temperatures.
authors.affiliation: Isermann Department of Chemical Engineering, Rensselaer Polytechnic Institute, Troy, NY 12180-3590.
authors.forename: T M
authors.initials: TM
authors.lastname: Przybycien
authors.forename: J P
authors.initials: JP
authors.lastname: Dunn
authors.forename: P
authors.initials: P
authors.lastname: Valax
authors.forename: G
authors.initials: G
authors.lastname: Georgiou
mesh_terms: D004926:Escherichia coli; D002479:Inclusion Bodies / Q000737:chemistry / Q000648:ultrastructure*; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry*; D013059:Spectrum Analysis, Raman; D001618:beta-Lactamases / Q000096:biosynthesis / Q000737:chemistry*
chemical_list: D011993:Recombinant Fusion Proteins; D001618:beta-Lactamases