Biology subjects

Basczok, M. M.

Publications and source records attributed to Basczok, M. M..

3 recordsLinked to original sources

bioRxiv2024

Different RNA recognition by ProQ and FinO depends on the sequence surrounding intrinsic terminator hairpins

Escherichia coli ProQ and FinO proteins both have RNA-binding FinO domains, which bind to intrinsic transcription terminators, but each protein recognizes distinct RNAs. To explore how ProQ and FinO discriminate between RNAs we transplanted sequences surrounding terminator hairpins between RNAs specific for each protein, and compared their binding to ProQ, the isolated FinO domain of ProQ (ProQNTD), and FinO. The results showed that the binding specificity of chimeric RNAs towards ProQ, ProQNTD, or FinO was determined by the origin of the transplanted sequence. Further analysis showed that the sequence surrounding the terminator hairpin, including a purine-purine mismatch, in natural RNA ligands of FinO and in chimeric RNAs weakened their binding by ProQNTD. Overall, our studies suggest that the discrimination between RNAs by ProQ and FinO is determined by RNA sequence elements surrounding the intrinsic terminator hairpin.

biochemistry↗

bioRxiv2024

RNA recognition by minimal ProQ from Neisseria meningitidis

Neisseria meningitidis minimal ProQ is a global RNA binding protein belonging to the family of FinO-domain proteins. The N. meningitidis ProQ consists only of the FinO domain accompanied by short N- and C-terminal extensions. To better understand how this minimal FinO-domain protein recognizes RNAs, we compared its binding to seven different natural RNA ligands of this protein. Next, two of these RNAs, rpmG-3 and AniS, were subject to further mutational studies. The data showed that N. meningitidis ProQ binds the lower part of the intrinsic transcription terminator hairpin, and that the single-stranded sequences on the 5 and 3' side of terminator stem are required for tight binding. However, the specific lengths of 5 and 3' RNA sequences required for optimal binding differed between the two RNAs. Additionally, our data show that the 2'-OH and 3'-OH groups of the 3' terminal ribose contribute to RNA binding by N. meningitidis ProQ. In summary, the minimal ProQ protein from N. meningitidis has generally similar requirements for RNA binding as the isolated FinO domains of other proteins of this family, but differs from them in detailed RNA features that are optimal for specific RNA recognition.

biochemistry↗

bioRxiv2020

Determinants of RNA recognition by the FinO domain of the Escherichia coli ProQ protein

The regulation of gene expression by small RNAs in Escherichia coli depends on RNA binding proteins Hfq and ProQ, which bind mostly distinct RNA pools. To understand how ProQ discriminates between RNA substrates, we compared its binding to six different RNA molecules. Full-length ProQ bound all six RNAs similarly, while the isolated N-terminal FinO domain (NTD) of ProQ specifically recognized RNAs with Rho-independent terminators. Analysis of malM 3-UTR mutants showed that tight RNA binding by the ProQ NTD required a terminator hairpin of at least two base pairs preceding an 3 oligoU tail of at least four uridine residues. Substitution of an A-rich sequence on the 5 side of the terminator to uridines strengthened the binding of several ProQ-specific RNAs to the Hfq protein, but not to the ProQ NTD. Substitution of the motif in the malM-3 and cspE-3 RNAs also conferred the ability to bind Hfq in E. coli cells, as measured using a three-hybrid assay. In summary, these data suggest that the ProQ NTD specifically recognizes 3 intrinsic terminators of RNA substrates, and that the discrimination between RNA ligands by E. coli ProQ and Hfq depends both on positive determinants for binding to ProQ and negative determinants against binding to Hfq.

molecular biology↗