# Protein Assistants of Small Ribosomal Subunit Biogenesis in Bacteria
## 2 30S Ribosomal Subunit Biogenesis Factors
### RimP  
In _E. coli_, the _rimP_ gene encoding the RimP protein is located in the same operon as _rbfA_[16]. In Gram-negative bacteria, _rimP_ knockout causes slow cell growth [16; 101]; whereas in Gram-positive _Streptococcus pneumoniae_, the deletion of _rimP_ is lethal [102], and in _Mycobacterium fortuitum_, RimP is required for survival under stress conditions [103]. RimP has not been identified in eukaryotes and archaea, but it is highly conserved in bacteria. RimP is a small globular protein (approximately 17 kDa in _E. coli_) that consists of two domains; each domain has an \(\alpha/\beta\)-fold (Figure 3_g_) [102; 104; 105]. The N-terminal domain contains two \(\alpha\)-helices and a three-stranded \(\beta\)-sheet, whereas the C-terminal part consists of a single \(\alpha\)-helix and a five-stranded \(\beta\)-sheet. The N-terminal domain of RimP is structurally similar to RbfA, and the C-terminal domain has an Sm fold that is characteristic of proteins belonging to Sm/Lsm superfamily The representative of this superfamily in bacteria is the Hfq protein (whose role in the ribosome biogenesis is addressed in the next section). Unlike the Hfq protein, which forms a hexameric ring, RimP exists in a monomeric form.  
Deleting _rimP_ in _E. coli_ caused a temperature-sensitive growth phenotype, which differentiated RimP from other 30S subunit biogenesis factors [16]. In addition, at higher temperatures, the _ArmP_ strain exhibited the typical ribosome biogenesis defects: a reduced amount of 70S ribosomes, an increase in the amount of free 30S and 50S ribosomal subunits, and accumulation of immature 16S rRNA. Interestingly, none of the known 30S maturation factors (RimM, RbfA, Era, KsgA, and YjeQ) suppressed the defects of the _ArmP_ strain [16]. Moreover, the N-terminal domain of RimP (similarly to RbfA) did not suppress the slow growth of a _rbfA_ null mutant [16]. This was probably due to different surface charges of the proteins, which were predominantly negative for the N-terminal domain RimP and positive for RbfA. Therefore, the RimP protein may act independently of the other assembly factors and is crucial at higher temperatures. Similarly, to Hfq, a robust Sm fold apparently preserves the native structure of RimP at high temperatures. It may be assumed that RimP is overexpressed under heat-shock conditions to maintain efficient ribosome biogenesis at higher temperatures.  
Cryo-EM analysis of pre-30S subunits accumulating in the _ArmP_ strain showed a wide range of 30S assembly intermediates from \"early\" particles with an almost missing cryo-EM density for the head to nearly mature 30S subunits with body, platform, and head domains and a distorted helix 44 [106]. The presence of intermediates with an unformed central pseudoknot and depletion of S5 and S12 proteins interacting with helices of central PK is a characteristic feature of _ArmP_ pre-30S particles. Quantitative MS studies on _E. coli_ revealed that RimP accelerated the binding of the S5 and S12 to the 16S rRNA in vitro [100]. It was suggested that RimP takes part in stabilization of the central PK at the early stages of the 30S subunit maturation, and this process may occur before the head domain assembly [106]. Cryo-EM analysis of complexes of RimP with mature 30S subunits simulating different assembly stages (inactive/active forms and in combination with different 30S assembly factors) revealed that RimP was bound to the t majority of the 30S assembly states (Figure 4f) [30]. This indicated that RimP (similarly to RbfA) is deeply involved in the 30S subunit maturation process and can assist in the formation of the central pseudoknot and later stages of the 30S assembly. RimP interacts with the helix 44 in the vicinity of KsgA-binding site on the 30S subunit, and apparently prevents A1492 and A1493 residues at the top of h44 from approaching S12, which maintains translation fidelity [30]. So, RimP, together with KsgA, delay h44 positioning at the 30S subunit interface at the late stage of the 30S assembly.