Structural Aspects of RbfA Action during Small Ribosomal Subunit Assembly

Datta, Partha P.; Wilson, Daniel N.; Kawazoe, Masahito; Swami, N.K.; Kaminishi, T.; Sharma, Manjuli R.; Booth, T.M.; Takemoto, Chie; Fucini, Paola; Yokoyama, Shigeyuki; Agrawal, Rajendra K.

doi:10.1016/j.molcel.2007.08.026

Cited by 92 publications

(133 citation statements)

References 61 publications

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“…A recent publication (Goto et al 2011) has reported a functional interplay between YjeQ and RbfA during maturation of the 30S subunit. RbfA is a small protein that binds the 30S subunit at the junction of the head and body, and its binding alters the position of helices 44 and 45 (Datta et al 2007). Interestingly, RbfA seems to promote the association of YjeQ to the 30S subunits, and binding of YjeQ stimulates the dissociation of RbfA from the small subunit (Goto et al 2011).…”

Section: Discussionmentioning

confidence: 99%

Understanding ribosome assembly: the structure of in vivo assembled immature 30S subunits revealed by cryo-electron microscopy

Jomaa¹,

Stewart²,

Martín‐Benito³

et al. 2011

RNA

111

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Four decades after early in vitro assembly studies demonstrated that ribosome assembly is a controlled process, our understanding of ribosome assembly is still incomplete. Just as structure determination has been so important to understanding ribosome function, so too will it be critical to sorting out the assembly process. Here, we used a viable deletion in the yjeQ gene, a recognized ribosome assembly factor, to isolate and structurally characterize immature 30S subunits assembled in vivo. These small ribosome subunits contained unprocessed 17S rRNA and lacked some late ribosomal proteins. Cryo-electron microscopy reconstructions revealed that the presence of precursor sequences in the rRNA induces a severe distortion in the 39 minor domain of the subunit involved in the decoding of mRNA and interaction with the large ribosome subunit. These findings suggest that rRNA processing events induce key local conformational changes directing the structure toward the mature assembly. We concluded that rRNA processing, folding, and the entry of tertiary r-proteins are interdependent events in the late stages of 30S subunit assembly. In addition, we demonstrate how studies of emerging assembly factors in ribosome biogenesis can help to elucidate the path of subunit assembly in vivo.

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Section: Discussionmentioning

confidence: 99%

Understanding ribosome assembly: the structure of in vivo assembled immature 30S subunits revealed by cryo-electron microscopy

Jomaa¹,

Stewart²,

Martín‐Benito³

et al. 2011

RNA

111

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“…16S RNA bridges B5 and B6 have intersubunit contacts on the 30S subunit contributed entirely by helix 44. Helix 44 has been shown to be organized late during the assembly of the 30S subunit and has been found to be disordered (18,19) or distorted (20)(21)(22) in structures of some of the assembly intermediates of the small subunit from various species.…”

Section: Significancementioning

confidence: 99%

Initial bridges between two ribosomal subunits are formed within 9.4 milliseconds, as studied by time-resolved cryo-EM

Shaikh

Yassin

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Significance The protein-synthesizing machinery of the cell, the ribosome, is made up of two subunits, which in bacteria are held together by 12 molecular bridges. Understanding the mechanism and timeline of bridge formation is crucial to understanding the mechanism of bacterial translation initiation. To study the timeline of bridge formation, we developed microfluidic devices that mix two interacting components, spray the mixture onto an electron microscopy grid, and flash freeze the grid for a minimum reaction time of 9.4 ms. This study shows for the first time to the authors' knowledge that eight of the 12 bridges form within 9.4 ms, whereas the remaining four bridges take longer than 43 ms to form.

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“…Structurally, it has been shown that YjeQ, RbfA, and Era all bind in close proximity to the decoding center of the mature 30S subunit (Sharma et al 2005;Datta et al 2007;Guo et al 2011;Jomaa et al 2011b), suggesting that these factors play a joint role in the organization of the functional core of the small subunit. Genetic evidence suggests that RimM binds to a distinct region of the head of the 30S subunit comprised of helices 31 and 33b of the 16S rRNA and r-proteins S13 and S19 right above the decoding center (Lovgren et al 2004).…”

Section: Introductionmentioning

confidence: 99%

Escherichia coli rimM and yjeQ null strains accumulate immature 30S subunits of similar structure and protein complement

Leong

Kent

Jomaa

et al. 2013

RNA

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Assembly of the Escherichia coli 30S ribosomal subunits proceeds through multiple parallel pathways. The protein factors RimM, YjeQ, RbfA, and Era work in conjunction to assist at the late stages of the maturation process of the small subunit. However, it is unclear how the functional interplay between these factors occurs in the context of multiple parallel pathways. To understand how these factors work together, we have characterized the immature 30S subunits that accumulate in ΔrimM cells and compared them with immature 30S subunits from a ΔyjeQ strain. The cryo-EM maps obtained from these particles showed that the densities representing helices 44 and 45 in the rRNA were partially missing, suggesting mobility of these motifs. These 30S subunits were also partially depleted in all tertiary ribosomal proteins, particularly those binding in the head domain. Using image classification, we identified four subpopulations of ΔrimM immature 30S subunits differing in the amount of missing density for helices 44 and 45, as well as the amount of density existing in these maps for the underrepresented proteins. The structural defects found in these immature subunits resembled those of the 30S subunits that accumulate in the ΔyjeQ strain. These findings are consistent with an "early convergency model" in which multiple parallel assembly pathways of the 30S subunit converge into a late assembly intermediate, as opposed to the mature state. Functionally related factors will bind to this intermediate to catalyze the last steps of maturation leading to the mature 30S subunit.

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Structural Aspects of RbfA Action during Small Ribosomal Subunit Assembly

Cited by 92 publications

References 61 publications

Understanding ribosome assembly: the structure of in vivo assembled immature 30S subunits revealed by cryo-electron microscopy

Understanding ribosome assembly: the structure of in vivo assembled immature 30S subunits revealed by cryo-electron microscopy

Initial bridges between two ribosomal subunits are formed within 9.4 milliseconds, as studied by time-resolved cryo-EM

Escherichia coli rimM and yjeQ null strains accumulate immature 30S subunits of similar structure and protein complement

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