Incorporation of dinitrophenyl derivatives of proteins S6, S13, S16, and S18 into the 30 S subunit of Escherichia coli ribosomes by total reconstitution.
This is the third paper in a series (Olah, T. V., Olson, H. M., Glitz, D. G., and Cooperman, B. S. (1988) J. Biol. Chem. 263, 4795-4800; Olson, H. M., Olah, T., Cooperman, B. S., and Glitz, D. G. (1988) J. Biol. Chem. 263, 4801-4806) describing the use of 2,4-dinitrophenyl (DNP) derivatives of Escherichia coli 30 S ribosomal proteins to locate the positions of these proteins within the 30 S subunit by immune electron microscopy. In it we describe the derivatization of proteins S6, S13, S16, and S18 with [3H]2,4-dinitrofluorobenzene, identify the nature of the derivatized amino acids within each protein, and demonstrate that each DNP protein, denoted DNP-Sx, can be taken up into a reconstituted 30 S subunit when added to a reconstitution mixture containing 16 S rRNA and total 30 S protein depleted in Sx. We further demonstrate that each DNP-Sx binds within the 30 S subunit in a position identical or similar to that of the unmodified Sx protein, as judged by its meeting one or more of the following three criteria: (i) unmodified Sx competes with the uptake of DNP-Sx into 30 S subunits; (ii) DNP-Sx restores functional activity to those single protein omission reconstitution particles lacking full activity; (iii) DNP-Sx induces the uptake of proteins into 30 S subunits that depend on the presence of Sx. The fourth paper in this series (Montesano-Roditis, L., McWilliams, R., Glitz, D. G., Olah, T. V., Perrault, A. R., and Cooperman, B. S. (1993) J. Biol. Chem. 268, 18701-18709), which follows this one, describes the localization of the DNP-Sx proteins within the 30 S subunit by immune electron microscopy.
Olah
T V
TV
Department of Chemistry, University of Pennsylvania, Philadelphia 19104-6323.
Perrault
A R
AR
Cooperman
B S
BS
Montesano-Roditis
L
L
McWilliams
R
R
Glitz
D G
DG
D000596:Amino Acids / Q000378:metabolism; D001667:Binding, Competitive; D004139:Dinitrofluorobenzene / Q000378:metabolism*; D004926:Escherichia coli / Q000378:metabolism*; D029968:Escherichia coli Proteins; D016253:Microscopy, Immunoelectron; D012329:RNA, Bacterial / Q000378:metabolism; D012336:RNA, Ribosomal, 16S / Q000378:metabolism; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*
D000596:Amino Acids; D029968:Escherichia coli Proteins; D012329:RNA, Bacterial; D012336:RNA, Ribosomal, 16S; D038601:Ribosomal Protein S6; D012269:Ribosomal Proteins; C023509:ribosomal protein S16; C041184:ribosomal protein S18; C512838:rpsM protein, E coli; D004139:Dinitrofluorobenzene