Induction of polynucleotide-protein cross-linkages by ultraviolet irradiation. Peculiarities of the high-intensity laser pulse irradiation.
The efficiency and specificity of RNA-protein cross-linking in the 30S subunit of Escherichia coli ribosomes, induced by low-intensity (10(15) photons cm-2 s-1, 254 nm) and high-intensity [(1.6-6.8) X 10(24) photons cm-2 s-1, 266 nm, pulse duration 10(-8) s] ultraviolet radiation, are studied. Under the former conditions proteins S4, S7 and S9/S11, and under the latter conditions these proteins together with S3, S18 and S20, are cross-linked to 16S RNA. Biphotonic processes operate in the latter case. In the presence of 2-mercaptoethanol cross-linking occurs either directly, via a higher excited state or via activated intermediates with life-times less than 25 ns. Cross-links thus formed are specific, i.e. they are formed between regions of macromolecules which are in contact in the native (non-disturbed) complex prior to excitation. The efficiency of cross-linking (per photon absorbed) is 20-100 times higher upon two-step excitation as compared with single-step excitation and an analysable number of cross-links are produced in a single pulse. Only base U-1239 of 16S RNA is cross-linked to protein S7 by low-intensity radiation, whereas the adjacent base, G-1240 is also involved in laser-induced cross-linking. A transition from the former to the latter conditions allows one to reduce the duration of irradiation from several minutes to several nanoseconds.
D001426:Bacterial Proteins / Q000528:radiation effects; D004926:Escherichia coli / Q000528:radiation effects; D007834:Lasers; D008433:Mathematics; D008623:Mercaptoethanol; D009698:Nucleoproteins / Q000528:radiation effects*; D011485:Protein Binding / Q000528:radiation effects; D012329:RNA, Bacterial / Q000528:radiation effects; D012335:RNA, Ribosomal / Q000528:radiation effects; D012269:Ribosomal Proteins / Q000528:radiation effects; D014466:Ultraviolet Rays
D001426:Bacterial Proteins; D009698:Nucleoproteins; D012329:RNA, Bacterial; D012335:RNA, Ribosomal; D012269:Ribosomal Proteins; D008623:Mercaptoethanol