Rrna METHYLTRANSFERASES and THEIR ROLE in RESISTANCE to ANTIBIOTICS Rrnk METILTRANSFERAZE I NJIHOVA ULOGA U REZISTENCIJI NA ANTIBIOTIKE
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J Med Biochem 2010; 29 (3) DOI: 10.2478/v10011-010-0030-y UDK 577.1 : 61 ISSN 1452-8258 J Med Biochem 29: 165 –174, 2010 Review article Pregledni ~lanak rRNA METHYLTRANSFERASES AND THEIR ROLE IN RESISTANCE TO ANTIBIOTICS rRNK METILTRANSFERAZE I NJIHOVA ULOGA U REZISTENCIJI NA ANTIBIOTIKE Ivana Mori}1, Miloje Savi}1,2, Tatjana Ili}-Tomi}1, Sandra Vojnovi}1, Sanja Bajki}1, Branka Vasiljevi}1 1Institute of Molecular Genetics and Genetic Engineering, University of Belgrade, Belgrade 2Department of Biochemistry, Emory University School of Medicine, Atlanta, USA Summary: Methyltransferases (MTases), a large protein Kratak sadr`aj: Metiltransferaze (MTaze), koje ~ine veliku superfamily, commonly use S-adenosyl-L-methio nine (SAM) proteinsku superfamiliju, kao donatora metil grupe naj~e {}e as the methyl group donor. SAM-dependant MTases koriste S-adenozil-L-metionin (SAM). SAM-zavisne MTaze methylate both nucleic acids (DNA, RNA) and proteins, metiluju nukleinske kiseline (DNK, RNK) i proteine, mo - and thus modulate their activity, function and folding. duli{u}i tako njihovu aktivnost, funkciju i strukturnu orga - Methylation of G1405 or A1408 nucleotides of 16S rRNA nizaciju. Metilacija G1405 ili A1408 baza u 16S rRNK in aminoglycoside-producing microorganisms confers the mikroorganizama koji proizvode aminoglikozide obezbe|u - resistance to their own toxic product(s). This mechanism of je rezistenciju na sopstvene toksi~ne proizvode. Ovaj meha - resistance has been considered as unique to antibiotics nizam rezistencije je donedavno bio opisan samo kod producers until recently. Since 2003, methylation of 16S proizvo|a~a antibiotika. Od 2003. godine i kod patogenih rRNA as a mechanism of resistance is increasingly bakterija bele`i se neprestan porast rezi sten cije na ami - emerging in pathogenic bacteria. This represents a major noglikozide putem ovog mehanizma, {to predstavlja veliku threat towards the usefulness of aminoglycosides in the pretnju efikasnoj upotrebi aminoglikozida u klini~koj praksi. clinical practice. A potential solution to the problem invol- Jedno od mogu}ih re{enja problema le`i u razvoju novih ves the design of novel compounds that would act against jedinjenja koja bi efikasno delovala na nova mesta u okviru new ribosomal targets. The second approach to the issue ribozoma. Drugi pristup re{avanju ovog problema uklju~uje includes the development of resistance MTases’ inhibitors, razvoj inhibitora MTaza odgovornih za rezisten ciju, sa with the idea to prevent them from modifying the bacterial idejom da se onemogu}i modifikacija bakterijske rRNK i na rRNA, and thus reinstate the therapeutic power of existing taj na~in vrati terapeutska efikasnost postoje}im aminogli- aminoglycosides. As the latter approach has considerable ko zidima. Fundamentalna istra`ivanja vezana za proteinsku potential, it is obvious that fundamental research related ekspresiju, potpuno razumevanje mehanizma rezistencije to protein expression, in-depth understanding of the kao i razre{enje tercijarne strukture proteina su neophodan mechanism of action and resolving a tertiary structure of preduslov za primenu inhibitora 16S rRNK MTaza u 16S rRNAs MTases are prerequisites for application in medicini. medicine. Klju~ne re~i: aminoglikozidi, metiltransferaze, rezisten - Keywords: aminoglycosides, methyltransferases, resis- cija, ribozom tan ce, ribosome Introduction Methyltransferases (MTases), which are a large, diverse and biologically important protein super - family, most commonly use the ubiquitous S-ade- Address for correspondence: nosyl-L-methio nine (SAM) molecule as a methyl Ivana Mori}, PhD group donor. SAM-dependant MTases methylate a Institute of Molecular Genetics and Genetic Engineering Vojvode Stepe 444a, P.O. Box 23, 11000 Belgrade, Serbia broad array of substrates – nucleic acids, proteins, Phone: +381 11 3976 034 polysaccharides, lipids, and a range of small mole- Fax: +381 11 3975 808 cules, after their primary synthesis, in both prokary - e-mail address: brmboznaªgmail.com otic and eukaryotic cells (1). Methylating nucleic 166 Mori} et al.: rRNA methyltransferases and resistance acids (DNA, RNAs) and proteins, MTases modulate It is noteworthy that for the proper reading of their activity, function and folding; hence, they guide the genetic code, post-transcriptional methylation of the cell’s fate. Methyl group transfer is an alkylation transport RNA (tRNA) that can occur at the bases reaction, and the atomic targets for the reaction can flanking the anticodon, and in some cases even be carbon, oxygen, nitrogen, sulfur or even halides within the anticodon itself, is important. It is believed (1–3). that modifications in positions flanking the anticodon affect the function of tRNA through relevant alter- The methylation of DNA directs bacterial destiny nation of the »rigidity« and »flexibility« of tRNA, which by controlling numerous processes, including chro- mo some replication, mismatch repair, transcriptional impinge directly on proper base pairing required for regulation, and regulation of transposition. One of codon recognition (1). Remarkably, it has been re- the main functions of DNA methylation in bacteria is cently reported that DNA MTase plays hitherto un to protect the cell from the effect of foreign DNA. known role(s) in the post-transcriptional control of Bacterial restriction modification systems discriminate gene expression (i.e. mRNA) of a transcriptional between endogenous and foreign DNA, which is not regu lator important for Salmonella enterica patho - protected by methylation (4). Furthermore, DNA genicity (13). methyl ation is crucially involved in the control of Being of fundamental importance for cell replication fidelity, a process that directs faithful viability, it is not surprising that many antibiotics target transmission of genetic material during cell division. ribosomes (14). The main contact sites for the anti- Repair system corrects mismatches that occur as biotics are on the rRNA, rather than on the ribosomal repli cation errors in the newly synthesized unmet h - protein components (15), which is consistent with the ylated strand, ensuring accurate DNA replication (5). In view that the rRNAs carry out the primary functions of addition, a growing number of reports suggest that the ribosome, while the r-proteins have supporting DNA methylation may be a versatile regulator of viru- roles (16, 17). Several classes of antibiotics, such as lence gene expression in pathogenic bacteria during aminoglycosides, tetracycline, macrolides, lincosami - the course of infection (6, 7). des and strepto gramin B, target the rRNA-rich surfa - Translation of the genetic information encoded ces on the 30S and 50S ribosomal subunits, inter- in a DNA molecule into a functional protein is exe- fering with their functions in protein synthesis (14). cuted by the ribosome. The ribosome is nature’s Not surprisingly, therefore, changes that confer anti- largest, most complex and most ancient enzyme that biotic resistance to these chemotherapeutics mainly consists, even in the simplest organisms, of more consist of nucleotide methylations or base substitu - than fifty different proteins (r-proteins) and three tions (15). ribosomal RNAs (5S, 16S and 23S rRNA in proka ry - otes). Preser vation of its function has been coupled with an overall conservation of structure, and ribo- Antibiotics, resistance and methylation somes in organisms as phylogenetically distinct as Actinomycetes, ubiquitous Gram-positive bacte- bacteria and archaea show a remarkable degree of ria, are prolific producers of antibiotics and other resemblance. Furthermore, numerous evidence indi - secondary metabolites. After the discovery of strepto- cates that eukaryotic ribosomes, including those in mycin and other potent antibiotics, isolated from soil human cells, are also similar in structure to their actinomycetes, antibiotics were defined as »chemi cal prokaryotic counterparts. This complex, dynamic ma- substances of microbial origin, that possess antibiotic cro molecular ribonucleoprotein machine translates, powers«, on the assumption that these com pounds in a multi-stage process, the genetic information were exhibiting their natural roles (18). Their bona encoded in messenger RNA (mRNA) into proteins fide environmental activities are not known, although (8). Correct assembly, that is subjected to intricate there is evidence that the majority of low-molecular- control and aided by a multitude of assembly factors, weight organic compounds made and secreted by is pre requisite for its proper functioning (9). While the microorganisms play roles as cell-signaling molecules biological significance of r-protein modifications by in the environment (19). methylation is poorly understood (10), the impor - tance of methylations of rRNAs that carry out ‘house- In order to avoid the toxic effects of antibiotics, keeping’ roles essential for the general functioning of antibiotic-producing microorganisms have developed rRNA, during protein synthesis, has been recognized. specific mechanisms to overcome the toxicity of their Bacterial rRNAs can contain over thirty ’house- own product(s). Several mechanisms of resistance to keeping’ modifications, all of which are added post- antibiotics have been recognized to be engaged in transcriptionally (11). The sites of ‘house-keeping’ produ cers’ protection, including target site modifi- methylations have been most accurately mapped in cation through mutation or modifications, e.g. rRNA Escherichia coli. Its ribosome