Structural studies of the Staphylococcus aureus ribosome Iskander Khusainov
To cite this version:
Iskander Khusainov. Structural studies of the Staphylococcus aureus ribosome. Biophysics. Université de Strasbourg, 2015. English. NNT : 2015STRAJ071. tel-01403855
HAL Id: tel-01403855 https://tel.archives-ouvertes.fr/tel-01403855 Submitted on 28 Nov 2016
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UNIVERSITÉ DE STRASBOURG
ÉCOLE DOCTORALE des Sciences de la Vie et de la Santé IGBMC - CNRS UMR 7104 - Inserm U 964
THÈSE présentée par : Iskander KHUSAINOV
soutenue le : 27 Novembre 2015
pour obtenir le grade de : !"#$%&'($')R%+,-$&.,# '($'0#&1.2!%&3 Discipline/ Spécialité : Biophysique et biologie structurale (Biophysics and Structural biology)
Structural studies of the Staphylococcus aureus ribosome Etudes structurales du ribosome de Staphylococcus aureus
THÈSE dirigée par : M. YUSUPOV Marat Directeur de recherche, Université de Strasbourg Mme. YUSUPOVA Gulnara Directeur de recherche, Université de Strasbourg
RAPPORTEURS : M. JENNER Lasse B. Associate Professor, Aarhus University (Denmark) M. MECHULAM Yves Directeur de recherche, École Polytechnique (Paris)
AUTRES MEMBRES DU JURY : M. GEISSMANN Thomas Chargé de recherches, ENS de Lyon (Lyon) M. MARZI Stefano Chargé de recherches, Université de Strasbourg
AcknowledgementsAcknowled ge ments
ACKNOWLEDGEMENTS
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DISCSSION 888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888 PERSPECTIES 88888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888 REFERENCES 88888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888 5 FiguresF ig ures and Tables TABLE OF FIGURES AND TABLES FIGRES7 F1$%`V 8 S `%H %`:C Q`$:J1<: 1QJ :JR `%JH 1QJ:C s1 Vs Q` .V `1GQsQIV 88888888888888888888888888888888888888888888 F1$%`V 8 CQI]Qs1 1QJ Q` G:H V`1:C :JR V%@:`7Q 1H `1GQsQIVs :JR .V HQIIQJ HQ`V5 G:sVR QJ 6R`:7 s %R1Vs88888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888888 F1$%`V 8 SH.VI: 1H R1:$`:I Q` G:H V`1:C ]`Q V1J s7J .Vs1s 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RXHQ%0`1` RVs HQI]QsXs :J 1H:JHX`V%6 ]Q VJ 1VCs8 LV `:0:1C : HQIIVJHX :0VH CXC:GQ`: 1QJ R% ]`Q QHQCV RV ]%`1`1H: 1QJ RV C: ]`Q X1JV J: 10V VEF % 1C1s:J HQIIV sQ%`HV C: ``:H 1QJ H7 QsQC1_%V C1G`V RVs `1GQsQIVs RVs HVCC%CVs HVL:8 L: ]%`V X5 C.QIQ$XJX1 X V C: s :G1C1 X RV C: ]`Q X1JV ]%`1`1XV : X X :J:C7sXV ]:` SDSRPAGE5 R1``%s1QJ R7J:I1_%V RV C: C%I1W`V5 R1``%s1QJ RV C: C%I1W`V I%C 1:J$CV5 Vss:1 RV RXH:C:$V .V`I1_%V V C: s]VH `QIX `1V RV I:ssV8 LVs ]`VI1V`s H`1s :%6 QJ X X QG VJ%s ]:` `VH.V`H.V `QGQ 1_%V ^F1$ I_8 L:J:C7sV ]:` R1```:H 1QJ RV HVs H`1s :%6 :% s7JH.`Q `QJ 2;V2 :0X`X Y `V RVs H`1s :%6 RV ]`Q X1JV8 CV]VJR:J 5 C: _%:C1 X RV C: R1```:H 1QJ JV s%``1 ]:s ; HQCCVH V` CVs RQJJXVs8 20 Résumé de thèse F1$ I8 LVs H`1s :%6 RV ]`Q X1JV VEF QG VJ%s ]:` `VH.V`H.V `QGQ 1_%V8 EJ Q% `V CQ] 1I1s: 1QJ RV C:%H`12 :CC12: 1QJ%R; VEF X :1 1I]Qss1GCV VJ `:1sQJ R% I:J_%V RV I: X`1VC8 L: ]%`1`1H: 1QJ RV C: ]`Q X1JV J: 10V `V_%1sV RV JQIG`V%sVs HVCC%CVs .%I:1JVs5 _%1 sQJ HQ( V%68 NQ%s :0QJs RQJH RXH1RX RV HCQJV` V V6]`1IV` C: ]`Q X1JV .X X`QCQ$%V8 IJ1 1:CVIVJ 5 H:` VCCV Q```V C: ]C%s `:H1CV5 C: IQ1Js H.W`V V CV ]C%s `:]1RV s7s WIV RV6]`Vss1QJ5 E8 HQC1 : X X sXCVH 1QJJXV8 CV]VJR:J 5 JQ%s :0QJs H.Q1s1 RV ]Q%`s%10`V ]:` C: ]%`1`1H: 1QJ RV C: ]`Q X1JV VEF `VHQIG1J:J V R:Js RVs G:H X`1Vs8 PQ%` CV IQIVJ 5 CV `:0:1C Vs ; CX :]V RV HCQJ:$V RV C: `X$1QJ HQR:J V RV C: ]`Q X1JV VEF R:Js %J 0VH V%` RV6]`Vss1QJ G:H X`1VJ ]ETR G ^H1sR :$5 C: .`QIG1JV s1 V RV HC10:$V5 `Xs1s :JHV ; C:I]1H1CC1JV5 IPTG 1JR%H 1GCV_8 L: $WJV HQI]CV RVEF .%I:1J Q% sVs ``:$IVJ s QJ X X :I]C1`1Xs VJ % 1C1s:J C: G1GC1Q .W_%V HVL: ADNH 1JHQ`]Q`XV R:Js %J G:H X`1Q].:$V8 LV `:0:1C Vs Q%=Q%`s VJ HQ%`s8 21 Abbreviations ABBREVIATIONS R1GQsQIV7 ::R RNA :I1JQ:H7CR RNA ARs1 V :I1JQ:H7C s1 V5 ^AR RNA T ARs1 V RNA_ EF VCQJ$: 1QJ `:H Q` ^VEF T V%@:`7Q 1H EF_ ERs1 V V61 s1 V ^ER RNA T ERs1 V RNA_ IF 1J1 1: 1QJ `:H Q` ^VIF T V%@:`7Q 1H IF_ IRNA IVssVJ$V` `1GQJ%HCV1H :H1R PTC ]V] 1R7C `:Js`V`:sV HVJ `V PRs1 V ]V] 1R7C s1 V5 ^PR RNA T PRs1 V RNA_ RF `VCV:sV `:H Q` ^VRF T V%@:`7Q 1H RF_ RP `1GQsQI:C ]`Q V1Js RRF `1GQsQIV `VH7HC1J$ `:H Q` `RNA `1GQsQI:C `1GQJ%HCV1H :H1R SD S.1JVRD:C$:`JQ RNA `:Js`V` `1GQJ%HCV1H :H1R MV .QRs7 NMR J%HCV:` I:$JV 1H `VsQJ:JHV C`7QREM H`7QRVCVH `QJ I1H`QsHQ]7 LCRMSLMS C1_%1R H.`QI: Q$`:].7 HQ%]CVR I:ss s]VH `QIV `7 DLS R7J:I1H C1$. sH: V`1J$ MALLS I%C 1:J$CV C:sV` C1$. sH: V`1J$ MALDI I: `16 :ss1s VR C:sV` RVsQ`] 1QJL1QJ1<: 1QJ PCR ]QC7IV`:sV H.:1J `V:H 1QJ PAGE ]QC7:H`7C:I1RV $VC VCVH `Q].Q`Vs1s S8 :%`V%s7 B8 s%G 1C1s B:H1CC%s s%G 1C1s E8 HQC1 EsH.V`1H.1: HQC1 MRSA IV .1H1CC1JR`Vs1s :J S8 :%`V%s I`MRSA I%C 1 `Vs1s :J MRSA MSSA IV .1H1CC1JRs%sHV] 1GCV S8 :%`V%s JI`MRSA JQJRI%C 1 `Vs1s :J MRSA PBPs ]VJ1H1CC1J G1JR1J$ ]`Q V1Js S8 :%`V%s S :].7CQHQHH%s :%`V %s T8 .V`IQ].1C%s T.V`I%s .V`IQ].1C%s TSS Q61H s.QH@ s7JR`QIV ISA 0:JHQI7H1JR1J V`IVR1: V S8 :%`V%s RSA 0:JHQI7H1JR`Vs1s :J S8 :%`V%s HO Q`CR HV:C . O`$:J1<: 1QJ 5A s1$I: `:H Q` A 5B s1$I: `:H Q` B C`7QRVCVH `QJ I1H`QsHQ]77 CMOS HQI]CVIVJ :`7 IV :CRQ61RV sVI1HQJR%H Q` 22 Abbreviations CTF HQJ `:s `:Js`V` `%JH 1QJ DDD R1`VH RV VH Q` RV01HV DQE RV VH 10V _%:J %I V``1H1VJH7 FEG `1VCR VI1ss1QJ $%J FSC FQ%`1V` s.VCC HQ``VC: 1QJ SNR s1$J:CR QRJQ1sV `: 1Q TEM `:JsI1ss1QJ VCVH `QJ I1H`QsHQ]7 RV:$VJ s7 DMSO R1IV .7C sQC`Q61RV DTT R1 .1Q .`V1 QC EDTA V .7CVJVR1:I1JV V `::HV 1H :H1R EG V .7CQVJV $C7HQC GDP $%:JQs1JV R1].Qs].: V GTP $%:JQs1JV `1].Qs].: V LB C7sQ$VJ7 G`Q . 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VR QJ `1$%`Vs ARE8 D T PQs1 1QJ Q` J%HCVQ 1RV A ^`VR_5 `16VR G7 .V CCARVJR ^GC%V_ Q` .V PR RNA GQ%JR Q .V B8 s%G 1C1s `1GQsQIV8 E T T.V J%HCVQ 1RV A 1s JQ 01s1GCV QJ .V I:] Q` S8 :%`V%s `1GQsQIV R%V Q 1 s .1$. `CV61G1C1 78 OJV Q` .V H.:`:H V`1s 1H `V: %`Vs Q` .V `1GQsQIVs ``QI IVsQ].1C1H G:H V`1: :`V .V V6 VJs1QJs Q` `RNAs8 A HQI]:`1sQJ Q` .V sVHQJR:`7 s `%H %`Vs Q` .V S :JR S `RNA 1:s ]V``Q`IVR %s1J$ HQI]:`: 10V RNA 1VG ^CR_ s1 V ^C:JJQJV V :C85 _8 FQ` .1s :J:C7s1s 1V H.QsV QJV IVsQ].1C1H G`:IR]Qs1 10V G:H V`1%I ^ S8 :%`V%s _5 QJV IVsQ].1C1H G`:IRJV$: 10V G:H V`1%I ^ E8 HQC1 _ G:H V`1:5 QJV .V`IQ].1C1H G`:IRJV$: 10V G:H V`1%I ^ T8 .V`IQ].1C%s _ :JR QJV :`H.:V: ^H:CQ:`H%C: I:`1sIQ` %1 _8 SV0V`:C `RNA .VC16Vs 1V`V `Q%JR Q GV V6 VJRVR 1J .V s `%H %`Vs Q` S8 :%`V%s :JR E8 HQC1 1J HQJ `:s Q T8 .V`IQ].1C%s :JR H8 I:`1sIQ` %1 8 SQIV Q` .VI5 :`V HQJsV`0VR 11 . 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Q` .V ]`Q V1Js s `QJ$C7 HQ``Vs]QJRs Q 1 s .VQ`V 1H:C 0:C%V8 T:GCV 8 M:ss s]VH `QIV `7 `Vs%C s Q` VEF ]`Q V1J s:I]CV Protein Score Covarage, % MW [kDa] Calc. pI MALDI results Elongation factor 2 (Homo sapiens) 154 60 95.2 6.41 LC MS/MS results Elongation factor 2 (Homo sapiens ) 15272.3 96.6 95.3 6.4 Gelsolin (Homo sapiens) 3883.8 63.6 81.4 5.5 Ribonucleoside-diphosphate reductase large subunit (Homo sapiens) 1712.9 46.5 90 6.9 Peptidyl-prolyl cis-trans isomerase FKBP4 (Homo sapiens) 936.8 38.3 51.8 5.2 Ribosomal protein S6 kinase alpha-3 (Homo sapiens) 41.5 1.5 80.5 6.7 C0712 **'8 2'-, -$ &3+ , #EFTT TQ $V H`7s :Cs Q` .%I:J VEF5 1Q s `: V$1Vs 1V`V H.QsVJ8 F1`s 5 1V sV:`H.VR `Q` H`7s :CC1<: 1QJ HQJR1 1QJs s1I1C:` Q .: ]%GC1s.VR `Q` 7V:s VEF5 8 M HV]Vs ]H 85 Q EG5 IM DTT5 8 IM N:N 5 T Q PEG ^JZ`$VJsVJ V :C85 _8 SVHQJR5 1V sV:`H.VR `Q` JV1 H`7s :CC1<: 1QJ HQJR1 1QJs %s1J$ :CC HQIIV`H1:C sH`VVJs ``QI SIGMA5 H:I] QJ RVsV:`H.5 MQCVH%C:` R1IVJs1QJs5 Q%1:$VJ7 CC:ss1Hs5 P`Q V1J HQI]CV65 N%HCV165 MPD5 JCSGU5 PEGs5 PEG 1QJ ]H5 IJRV65 AJ1QJs5 C: 1QJs5 S:C R5 1<:`R I :JR 1<:`R II :0:1C:GCV : .V IGBMC H`7s :CC1<: 1QJ `:H1C1 78 C`7s :CC1<: 1QJ V6]V`1IVJ s 1V`V HQJR%H VR : C5 JC Q` ]`Q V1J 1J G%``V` III 1V`V I16VR 11 . 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T J%HCVQ 1RVs `C:J@1J$ .V $VJV :JR s16 J%HCVQ 1RVs Q` s]VH1`1H `Vs `1H 1QJ s1 Vs C1J@VR 11 . 1Q Q` .`VV J%HCVQ 1RV C1J@V`s8 T:GCV C1s s .V ]`1IV`s %sVR 1J .1s s %R78 T:GCV 8 P`1IV`s %sVR `Q` :I]C1`1H: 1QJ Q` .%I:J VEF8 RVs `1H 1QJ s1 Vs :`V .1$.C1$. VR 1J `VR5 C1J@V`s :`V .1$.C1$. VR 1J GC%V8 N:IV SV_%VJHV5 T F:$IVJ CVJ$ . F GCGC CATATG GGA ATGGTGAACTTCACGGTAGA 8 @G R GGCAGGTCAGCCGGGTTCTTGG F GCGTCAGCCCTGTTGTCAGAGTGG 8 @G R GCGC GGATCC TA CTACAATTTGTCCAGGAAGTTG Results F1$%`V 8 T1Q s `: V$1Vs Q` VEF HCQJ1J$ V6]V`1IVJ 8 PCR `V:H 1QJs 1V`V ]`V]:`VR :s `QCCQ1s5 1J : `1J:C 0QC%IV Q` -C7 DNA7 8 -C ^HVL: HDNA C1G`:`7_ PQC7IV`:sV G%``V`7 -C ^_ RNTP I167 -C ^8 IM s QH@ HQJHVJ `: 1QJ_ P`1IV` `Q`1:`R7 8 -C ^ -M s QH@_ P`1IV` `V0V`sV7 8 -C ^ -M s QH@_ Q H1$. 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GUC s :G1C1 Discussion 1J0QC0VR I:7 0:`78 SQIV G:H V`1: ^s%H. :s E8 HQC1 5 P8 H:`Q Q0Q`%I 5 S8 7].1I%`1%I :JR K8 ]JV%IQJ1: _ V6]`Vss .`VV R1``V`VJ ]`Q V1Js7 `1A5 RMF ^`1GQsQIV IQR%C: 1QJ `:H Q`_ :JR s.Q` HPF8 CQQ]V`: 10V G1JR1J$ Q` RMF :JR s.Q` HPF ]`Q V1Js Q .V `1GQsQIVs CV:Rs Q .V1` R1IV`1<: 1QJ5 1.V`V:s G1JR1J$ Q` `1A 1J.1G1 s `1GQsQIV :H 101 7 11 .Q% R1IV`1<: 1QJ8 T.V Q .V` $`Q%] Q` G:H V`1: 1JHC%R1J$ S8 :%`V%s 5 T8 .V`IQ].1C%s 5 L8 :H1RQ].1C%s H:``7 QJC7 QJV ]`Q V1J5 CQJ$ HPF8 T.V CQJ$ HPF ]`Q V1J HQIG1JVs .V ]`Q]V` 1Vs Q` :CC .`VV ]`Q V1Js8 I H:J R1IV`1 PerspectivesPer sp ectives PERSPECTIVES T.V 1Q`@ RVsH`1GVR 1J .1s .Vs1s 1:s .V `1`s s V] 1J .V HQJ V6 Q` `% %`V RV0VCQ]IVJ Q` R`%$s :$:1Js S :].7CQHQHH%s :%`V%s 8 OG :1JVR s `%H %`V Q` S8 :%`V%s `1GQsQIV 1s .V `1`s .1$.R `VsQC% 1QJ `1GQsQIV s `%H %`V ``QI ]: .Q$VJ1H G:H V`1: QG :1JVR %s1J$ H`7QREM8 T.V IQRVCC1J$ s V] 1s JQ ]V``Q`IVR 7V 5 .V`V`Q`V 1 1s JQ 1JHC%RVR 1J .1s .Vs1s I:J%sH`1] 8 OG :1JVR IQRVC Q` S8 :%`V%s 0:H:J `1GQsQIV 11CC GV %sVR C: V` QJ `Q` sQC01J$ s `%H %`Vs Q` .1s `1GQsQIV 1J HQI]CV6 11 . 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VCQJ$: 1QJ `:H Q` G `:]]VR 1J .V ]`V `:JsCQH: 1QJ s : V8 P`QH8 N: C8 AH:R8 SH18 8 S8 A8 5 T 8 B`QRV`sVJ5 D8E85 CCVIQJs5 8M85 C:` V`5 A8P85 MQ`$:JR:``VJ5 R8J85 1IGV`C75 B8T85 :JR R:I:@`1s.J:J5 8 ^_8 T.V s `%H %`:C G:s1s `Q` .V :H 1QJ Q` .V :J 1G1Q 1Hs V `:H7HC1JV5 ]:H :I7H1J5 :JR .7$`QI7H1J B5 QJ .V S `1GQsQI:C s%G%J1 8 CVCC 5 T 8 B`QJJV`5 S85 MQJ V1C5 H85 :JR P`X0Qs 5 G8 ^_8 RV$%C: 1QJ Q` 01`%CVJHV RV V`I1J:J s 1J S :].7CQHQHH%s :%`V%s7 CQI]CV61 7 :JR :]]C1H: 1QJs8 FEMS M1H`QG1QC8 RV08 5 T8 B`Q1J5 A85 AI%J s5 A85 B:15 85 S%$1IQ Q5 85 ER1:`Rs5 P8C85 M%`s.%RQ05 G85 SH.V`Vs5 S8H885 :JR R:I:@`1s.J:J5 8 ^_8 S `%H %`V Q` .V C:`$V `1GQsQI:C s%G%J1 ``QI .%I:J I1 QH.QJR`1:8 SH18 5 T 8 B%[email protected] T85 G1VsVG`VH. 5 J85 AC I:J5 R8B85 M%J`Q5 J8B85 M1VC@V5 T85 N1V`.:%s5 K8H85 BC:JH.:`R5 S8C85 :JR S]:.J5 C8M8T8 ^_8 S `%H %`V :JR R7J:I1Hs Q` .V I:II:C1:J `1GQsQI:C ]`V `:JsCQH: 1QJ HQI]CV68 MQC8 CVCC 5 T8 B%[email protected] T8 85 G1VsVG`VH. 5 J85 BV.`I:JJ5 E85 LQV`@V5 J85 R:I`: .5 D8J8F85 M1VC@V5 T85 IsIV`5 J85 H1CRVG`:JR5 P885 T%J$5 C8S85 N1V`.:%s5 K8H85 V :C8 ^_8 RV$%C: 1QJ Q` .V I:II:C1:J VCQJ$: 1QJ H7HCV G7 s%G%J1 `QCC1J$7 A V%@:`7Q 1HRs]VH1`1H `1GQsQIV `V:``:J$VIVJ 8 CVCC 5 T 8 C:C0Q`1 C5 F`QJ :C1 L5 LVQJ1 L5 T8G8 ^ _8 E``VH Q` `1`:I7H1J QJ ]`Q V1J s7J .Vs1s8 N: %`V 5 T 8 C:I]GVCC5 M8G85 C.VJ$5 A85 B`1CQ 5 A8F85 MQVCCV`5 A85 L7%I@1s5 D85 VVsCV`5 D85 P:J5 J85 H:``1sQJ5 S8C85 PQ V`5 C8S85 C:``:$.V`5 B85 V :C8 ^_8 MQ01Vs Q` 1HVRVIGVRRVR ]:` 1HCVs VJ.:JHV `VsQC% 1QJ 1J VCVH `QJ H`7QRI1H`QsHQ]78 S `%H %`V 5 T 8 <:JJQJV5'=8=85'?%G`:I:J1:J5'?85'?H.J:`V5'98B85' References BMC B1Q1J`Q`I: 1Hs 5 8 C.:IGV`s5 H8F8 ^ _8 MV .1H1CC1J `Vs1s :JHV 1J s :].7CQHQHH17 IQCVH%C:` :JR G1QH.VI1H:C G:s1s :JR HC1J1H:C 1I]C1H: 1QJs8 CC1J8 M1H`QG1QC8 RV08 5 T 8 C.:J5 P8F85 :JR FQs V`5 S8J8 ^ _8 RQCV Q` S:`A 1J 01`%CVJHV RV V`I1J:J ]`QR%H 1QJ :JR VJ01`QJIVJ :C s1$J:C `:JsR%H 1QJ 1J S :].7CQHQHH%s :%`V%s8 J8 B:H V`1QC8 5 T 8 C.:7VJ5 N8E85 S V1:` 5 P8D8S85 :JR BCQ15 D8M8 ^ _8 M1H`QG: H. 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Q` s1J$CV ]`Q V1J H`7s :Cs G7 IV:Js Q` V_%1C1G`1%I R1:C7s1s8 A`H.8 B1QH.VI8 B1Q].7s8 5 T 8 .:J$5 85 D%J@CV5 J8A85 :JR C: V5 J8H8D8 ^ _8 S `%H %`Vs Q` .V `1GQsQIV 1J 1J V`IVR1: V s : Vs Q` `: H.V 1J$8 SH1VJHV 5 T 8 .Q%5 J85 KQ`Qs VCV05 A85 L:JH:s V`5 L85 :JR NQCCV`5 H8F8 ^_8 C`7s :C s `%H %`Vs Q` S `1GQsQIVs GQ%JR Q `VCV:sV `:H Q`s RF5 RF :JR RF8 C%``8 O]1J8 S `%H 8 B1QC8 5 T 8 .Q%5 J85 L:JH:s V`5 L85 DQJQ.%V5 J8P85 :JR NQCCV`5 H8F8 ^_8 C`7s :C s `%H %`Vs Q` EFRGR `1GQsQIV HQI]CV6Vs `:]]VR 1J 1J V`IVR1: V s : Vs Q` `:JsCQH: 1QJ8 SH1VJHV 5 8 .Q%5 J85 L:JH:s V`5 L85 DQJQ.%V5 J8P85 :JR NQCCV`5 H8F8 ^_8 HQ1 .V `1GQsQIV .:JRs .V AR s1 V RNA Q .V P s1 V R%`1J$ EFRG TH: :C7 Iskander KHUSAINOV Structural studies of the Staphylococcus aureus ribosome Résumé Le ribosome est une machinerie cellulaire importante impliquée dans la synthèse protéique de toute cellule vivante. Par conséquent, le ribosome est l'une des principales cibles des antibiotiques naturels, qui sont capables de tuer les cellules bactériennes en bloquant la synthèse protéique. Toutefois, certaines bactéries sont résistantes à ces antibiotiques en raison de petites modifications au niveau de leurs ribosomes. Entre autres, Staphylococcus aureus ( S. aureus ) est un agent pathogène responsable de n !"#$%&$&'()*$+,( )&'-#./$&'+0$1'2R4 !!$5'6$&'&,#%+,%#$&'+#(&,.22()$&'78.),("( ,(9%$&' en complexe avec des ribosomes de bactéries non-résistantes, non-pathogènes, Gram négatives ont fourni un aperçu sans précédent des mécanismes d'action de ces antibiotiques. Cependant, aucune structure de ribosome de bactéries pathogènes, hautement résistantes, Gram positives telles que S. aureus )R.'$)+ #$' , '(7$),(*( $5 Dans cette étude, nous présentons la première structure de ribosome de S. aureus à haute résolution (3.9 Å) résolue par cryo-microscopie électronique (cryo-ME). Nous mettons en évidence plusieurs caractéristiques de l'organisation des ribosomes spécifiques des bactéries Gram-positives. Nous décrivons également le protocole de purification et de cristallisation du ribosome de S. aureus pour de futures études de cryo-ME et de cristallographie aux rayons X. ; %&'2$&'# &%2,.,&' ",$)%&'7.)&'+$&',#./.%<='*.+(2(,$# ),'2.'7$&+#(>,( )''2R +0$22$'., !(9%$'7%' ribosome de S. aureus et ses complexes fonctionnels dans un futur proche. La combinaison des méthodes de cristallographie aux rayons X et de cryo-ME aidera à atteindre cet objectif. Les résultats obtenus serviront de base pour le développement de nouveaux composés contre la bactérie pathogène et extrêmement # &(&,.),$'9%R$&,' S. aureus . Mots clés: Staphylococcus aureus , traduction, ribosome, structure, cristallographie, cryo-ME Summary The ribosome is a large cellular machinery that performs the protein synthesis in every living cell. Therefore, the ribosome is one of the major targets of naturally produced antibiotics, which can kill bacterial cells by blocking protein synthesis. However, some bacteria are resistant to these antibiotics due to small modifications of their ribosomes. Among them, Staphylococcus aureus ( S. aureus ), a severe pathogen that causes numerous infections in humans. The crystal structures of complexes of antibiotics with ribosomes from Gram-negative non-pathogenic non-resistant bacteria have provided unparalleled insight into mechanisms of antibiotics action and promoted the development of new semi- synthetic and synthetic antibiotics. However, the structure of the ribosome from Gram-positive pathogenic and highly resistant bacteria such as S. aureus was still unidentified. In this study we present the first high resolution structure of the ribosome from S. aureus solved at 3.9 Å by cryo-electron microscopy (cryo-EM). We demonstrate several features of the ribosome organization which are unique for Gram-positive bacteria. We also describe the protocol of purification and crystallization of S. aureus ribosome for future cryo-EM and X-ray crystallography studies. All the results obtained in this work will help to describe S. aureus ribosome and its functional complexes at the atomic level in the nearest future. The combination of X-ray crystallography and cryo- EM methods will help to achieve this aim. The obtained results will provide a foundation for the development of new compounds against pathogenic and extremely resistant bacteria S. aureus . Key words: Staphylococcus aureus , translation, ribosome, structure, crystallography, cryo-EM