Intrinsically Disordered C-Terminal Tails of E. coli Single-Stranded DNA Binding Protein Regulate Cooperative Binding to Single-Stranded DNA

Abstract

The homotetrameric E. coli single stranded DNA binding (SSB) protein plays a central role in DNA replication, repair and recombination. E. coli SSB can bind to long single-stranded (ss) DNA in multiple binding modes using all four subunits ((SSB)65 mode) or only two subunits ((SSB)35 binding mode), with the binding mode preference regulated by salt concentration and SSB binding density. These binding modes display very different ssDNA binding properties with the (SSB)35 mode displaying highly cooperative binding to ssDNA. SSB tetramers also bind an array of partner proteins, recruiting them to their sites of action. This is achieved through interactions with the last 9 amino acids (acidic tip) of the intrinsically disordered linkers (IDLs) within the four C-terminal tails connected to the ssDNA binding domains. Here we show that the amino acid composition and length of the IDL affects the ssDNA binding mode preferences of SSB protein. Surprisingly the number of IDLs and the lengths of individual IDLs together with the acidic tip contribute to highly cooperative binding in the (SSB)35 binding mode. Hydrodynamic studies and atomistic simulations suggest that the E. coli SSB IDLs show a preference for forming an ensemble of globular conformations, whereas the IDL from Plasmodium falciparum SSB forms an ensemble of more extended random coils. The more globular conformations correlate with cooperative binding.

Relating sequence encoded information to form and function of intrinsically disordered proteins

“…C-terminal disordered tails of E.coli single stranded DNA binding proteins belong to region R1 and these tails engender positive cooperativity in single stranded DNA binding. Cooperativity in single stranded DNA binding is abolished if the tails are eliminated or replaced with sequences drawn from the R3 region [63]. …”

“…This should be helpful for establishing the connections between changes to SCRs and functions / phenotypes controlled by polyampholytic sequences drawn from regions R2 and R3. A range of targets for such design efforts is readily available from the rich literature on IDPs with established functional roles for polyampholytic sequences [8,9,14,19,63,74–77]. Of course, the patterning of oppositely charged residues quantified by κ is not the only way to conceive of modulating SCRs.…”

Identification of inhibitors for single-stranded DNA-binding proteins in eubacteria

“…These differences are possibly due to intrinsic differences in binding activity, as well as a decrease in cooperative binding due to the truncation of nine amino acids from the C-termini of B. anthracis and S. aureus SSBAs. 26 The binding activities of all four SSBAs were considered qualified for EMSA and HTS assays. Glanzer et al…”

Variations in amino acid composition in bacterial single stranded DNA–binding proteins correlate with GC content

“…In the proposed model the Ct tail is bound to the OB fold in the absence of ssDNA, while in the presence of ssDNA the Ct tail is released, thus leaving it free for the interactions with other cellular proteins. Recently it was demonstrated that an intrinsically disordered C-terminal region of E. coli SSB protein participated in cooperative binding to ssDNA (16). All these results suggest an important regulatory role of Ct domain.…”

“…Recently, it was shown that shortening of C-terminal region of E. coli SSB protein had an impact on cooperative binding to ssDNA (16). It was also reported that extension of Ct domain slowed cell growth rate, indicating impaired protein function in vivo (14).…”