2. Special features of i-tRNA
Organisms possess two distinct methionine tRNAs, the i-tRNA and
elongator tRNAMet. Both the tRNAs possess CAU
anticodons. The i-tRNA decodes the initiation/start codons (AUG or a
related codon), and elongator tRNAMet decodes the AUG
codons found at internal positions in ORF. Like other tRNAs, these tRNAs
possess typical clover-leaf secondary structures. However, i-tRNAs
harbour at least three distinct structural features that distinguish
them from the elongator tRNAs (Fig. 2A ). Firstly, there exists
a Watson-Crick mismatch between nucleotides 1 and 72 (C1x A72 in E. coli ) at the top of the acceptor stem
(in contrast to the Watson-Crick base pairs found in elongator tRNAs at
this position). The 1x72 pair mismatch is crucial for, (i) recognition
of i-tRNA by Fmt (along with G2-C71,
C3-G70, and
A11-U24 base pairs) for formylation of
the amino acid attached to it; (ii) prevention of fMet-i-tRNA (a form of
peptidyl-tRNA) from hydrolysis by peptidyl-tRNA hydrolase (Pth), whose
role is to recycle tRNAs from peptidyl-tRNAs (Das and Varshney 2006);
and (iii) avoidance of its binding to EFTu and the ambiguity of its
participation at the step of elongation (Dutka et al. 1993; Guillon et
al. 1992; Hansen et al. 1986; Lee et al. 1992; Lee, Seong, and
RajBhandary 1991; Ramesh et al. 1997; Schulman and Pelka 1975; Seong,
Lee, and RajBhandary 1989; Seong and RajBhandary 1987; Varshney et al.
1991). Secondly, the bacterial i-tRNAs possess a
Pu11-Py24 base pair (unlike
Py11-Pu24 base pair in elongator tRNAs).
This position has been shown to impact formylation of the amino acid
attached to i-tRNA (Lee et al. 1991). Thirdly, all i-tRNAs possess a
highly conserved feature of three consecutive GC base pairs
(G29-C41,
G30-C40,
G31-C39, also stated as GC/GC/GC or 3GC
pairs) in the anticodon stem (Fig. 2A, i ), which enable its
direct binding to the P-site (Mandal et al. 1996; Marck and Grosjean
2002; Mayer et al. 2001; Varshney, Lee, and RajBhandary 1993). These
special features of i-tRNA not only distinguish it from elongator
tRNAMet (Fig. 2A) but also, along with IFs,
allow it to outcompete elongator tRNAs from binding into the P-site.
While the 3GC pairs in i-tRNA anticodon stem are highly conserved in all
domains of life, the first and third GC pairs
(G29-C41, and
G31-C39) have been found to be replaced
by A-U or G-U pairs in organisms such as mycoplasma, and
α-proteobacterial species (Ayyub et al. 2018; Dong et al. 2014; Samhita,
Shetty, and Varshney 2012). The middle one
(G30-C40) is the most crucial GC pair in
the bacterial i-tRNAs. In addition, it was shown that the first and the
third GC pairs are dispensable even in E. coli (Samhita et al.
2012). In fact, even the second G-C pair could be changed to a G-U pair
in some sequence contexts (Samhita et al. 2012; Shetty et al. 2017).
Although it should also be said that the mutations in the 3GC pairs do
not support a healthy growth of E. coli (Shetty et al. 2017).