Post-transcriptional control of gene expression: bacterial mRNA degradation

Many biological processes cannot be fully understood without detailed knowledge of RNA metabolism. The continuous breakdown and resynthesis of prokaryotic mRNA permit rapid production of new kinds of proteins. In this way, mRNA levels can regulate protein synthesis and cellular growth. Analysing mRNA degradation in prokaryotes has been particularly difficult because most mRNA undergo rapid exponential decay. Prokaryotic mRNAs differ in their susceptibility to degradation by endonucleases and exonucleases, possibly because of variation in their sequencing and structure. In spite of numerous studies, details of mRNA degradation are still largely unknown. This review highlights those aspects of mRNA metabolism which seem most influential in the regulation of gene expression.     

Pyridine Derivatives in Peppermint Oil

Five kinds of pyridine derivatives (1~5), including a novel compound, 5-phenyl-2-propylpyridine (4), were newly identified in peppermint oil.     

Chemical ribonucleases: VII. Effect of positively charged RNA-binding domains and hydrophobic fragments of the conjugates based on 1,4-diazabicyclo[2.2.2]octane and imidazole on their ribonuclease activity

The effect of the total positive charge in the RNA-binding domain of chemical ribonucleases that are conjugates of bisquaternary salts of 1,4-diazabicylo[2.2.2]octane and imidazole on the cleavage of an HIV-1 RNA fragment was studied. An increase in the positive charge from +2 to +4 was shown to result in a significant growth in the ribonuclease activity. Possible mechanisms of the interactions between structural moieties of chemical ribonucleases and RNA that enable an effective catalysis of the cleavage of phosphodiester bonds are discussed.     

Kinetic Parameters of Cleavage of CpA and UpA Sequences in an Oligoribonucleotide by Compounds Functionally Mimicking Ribonuclease A

Kinetic parameters of cleavage of CpA and UpA sites in an oligoribonucleotide under the action of artificial ribonuclease ABL3C1 were measured. The compounds were built of RNA-binding domain B, catalytic fragment C, linker L3 comprising 3 methylene groups, and aliphatic fragment A. The rate of cleavage of phosphodiester bonds in the CpA site within decaribonucleotide UUCAUGUAAA was shown to be 3.4 ± 0.2 times higher than in UpA. The rate of cleavage of phosphodiester bonds was found to depend on substrate length: a thousandfold increase in cleavage rate constant was observed for the CpA site in decaribonucleotide as compared with diribonucleoside monophosphate CpA. A slight decrease in the cleavage rates was observed for the reactions proceeding in different buffers at pH 7.0: imidazole > HEPES > phosphate > cacodylate. At the same time, the ratio of cleavage rates for CpA and UpA sites remained constant.     

Global and local motions in ribonuclease A: a molecular dynamics study

The understanding of protein dynamics is one of the major goals of structural biology. A direct link between protein dynamics and function has been provided by x-ray studies performed on ribonuclease A (RNase A) (B. F. Rasmussen et al., Nature, 1992, Vol. 357, pp. 423-424; L. Vitagliano et al., Proteins: Structure, Function, and Genetics, 2002, Vol. 46, pp. 97-104). Here we report a 3 ns molecular dynamics simulation of RNase A in water aimed at characterizing the dynamical behavior of the enzyme. The analysis of local and global motions provides interesting insight on the dynamics/function relationship of RNase A. In agreement with previous crystallographic reports, the present study confirms that the RNase A active site is constituted by rigid (His12, Asn44, Thr45) and flexible (Lys41, Asp83, His119, Asp121) residues. The analysis of the global motions, performed using essential dynamics, shows that the two beta-sheet regions of RNase A move coherently in opposite directions, thus modifying solvent accessibility of the active site, and that the mixed alpha/3(10)-helix (residues 50-60) behaves as a mechanical hinge during the breathing motion of the protein. These data demonstrate that this motion, essential for RNase A substrate binding and release, is an intrinsic dynamical property of the ligand-free enzyme.     

Population shift vs induced fit: the case of bovine seminal ribonuclease swapping dimer

Bovine seminal ribonuclease (BS-RNase) is a unique member of the pancreatic-like ribonuclease superfamily. This enzyme exists as two conformational isomers with distinctive biological properties. The structure of the major isomer is characterized by the swapping of the N-terminal segment (MxM BS-RNase). In this article, the crystal structures of the ligand-free MxM BS-RNase and its complex with 2'-deoxycitidylyl(3',5')-2'-deoxyadenosine derived from isomorphous crystals have been refined. Interestingly, the comparison between this novel ligand-free form and the previously published sulfate-bound structure reveals significant differences. In particular, the ligand-free MxM BS-RNase is closer to the structure of MxM BS-RNase productive complexes than to the sulfate-bound form. These results reveal that MxM BS-RNase presents a remarkable flexibility, despite the structural constraints of the interchain disulfide bridges and the swapping of the N-terminal helices. These findings have important implications to the ligand binding mechanism of MxM BS-RNase. Indeed, a population shift rather than a substrate-induced conformational transition may occur in the MxM BS-RNase ligand binding process.     

Conformational exchange divergence along the evolutionary pathway of eosinophil-associated ribonucleases

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Structure–activity relationships in new polycationic molecules based on two 1,4-diazabicyclo[2.2.2]octanes as artificial ribonucleases

In the present study, we designed and synthesised new polycationic molecules based on two 1,4-diazabicyclo[2.2.2]octane (DABCO) moieties with hydrophobic groups connected by different linkers. The structure and the RNA-cleavage activity relationships of this novel series of artificial ribonucleases (aRNases) were investigated.     

Structure–Function Study of Recombinant Onconase Variants

A method for expression of an onconase gene leading to a soluble form of the protein was developed. The enzymatic and cytotoxic properties of the protein's recombinant forms were studied. Recombinant onconase with an additional N-terminal Met residue isolated in non-denaturing conditions did not substantially differ from the native enzyme in ribonucleolytic activity. The addition of a 33-mer peptide containing auxiliary elements for the simplification of isolation and detection of the recombinant protein did not affect the enzyme properties of onconase. The method proposed is useful for the onconase structure–function relation studies and enables construction of onconase-based fusion proteins for anticancer therapy.     

Ribonuclease inhibitors

RNases are important enzymes of cell metabolism, influencing gene expression, affecting cell growth and differentiation, and participating in cell defense against pathogens and induction of apoptosis. Since RNases mostly occur in complex with their inhibitors in the cell, the inhibitors also play a role in the above processes. The review considers natural protein RNase inhibitors of animals, plants, and bacteria, as well as synthetic low-molecular-weight inhibitors. Special emphasis is placed on the prospective use of RNase inhibitors in the therapy of cancer and allergy. While RNases are widespread, the number of the available natural and synthetic inhibitors is limited. A pressing problem is to design highly effective low-molecular-weight inhibitors of the RNase activity of angiogenin and eosinophil-associated RNases for anticancer and antiallergy therapy.