BIOINORGANIC CHEMISTRY A Short Course Second Edition

MAGNESIUM AND CATALYTIC RNA 245

Tetrahymena pre - rRNA, beginning in 1982. Subsequent work showed that a
specifi c guanosine binding site (the G site) is found on the intron RNA, an
exogenous (from solution) guanosine (G OH ) binds there, and the intron pro-
vides an “ internal guide sequence ” (IGS) to bring the two exons (3 ′ exon
region and 5 ′ exon region) close together. (See Figure 6.1 .) In these processes,
the exon is considered to be the reaction substrate. In the fi rst of two trans-
esterifi cation reactions, G OH attacks at the 5 ′ splice site, becoming covalently
attached to the 5 ′ end of the intron. Subsequently, there is a conformational
change and the ribozyme ’ s 3 ′ - terminal guanosine (called the ω G) binds at the
“ G site. ” This positions the 3 ′ splice site for nucleophilic attack by the 3 ′ OH
of the 5 ′ exon; and in the second transesterifi cation the two exons are joined
and released, and the intron is liberated. After release of the ligated exons, the
intron ’ s 5 ′ region folds back to the active site and a third nucleophilic attack
(byω G) takes place to produce a circularized intron. At the same time an
RNA oligonucleotide is released carrying the exogenous G OH.^24 This splicing
mechanism has become known as the G - dependent mechanism. (See Figure
6.1 and Figure 6.3 .) Figure 6.3 shows the cleavage phosphotransesterifi ca-
tion — labeled chem.1 in Figure 6.1. The secondary structure of an entire group
I intron is shown in Figure 6.4 , and a close - up view of a transesterifi cation step
is illustrated in Figure 6.8 for PDB: 1ZZN.
Group I intron phosphotransesterifi cation reactions are carried out by a
conserved active site that contains a set of imperfect double helices named P1
through P9. (See Figure 6.4 .) P1 – P9 helices are organized into three domains:
P1 – P2, P4 – P6, and P3 – P9. Specifi cally, the Tetrahymena thermophila intron
contains two sets of coaxially stacked helices that overlap to create the active
site. These helices reside in two domains of approximately equal size: P4 – P6
and P3 – P9. P domains are defi ned as base - paired regions, whereas J domains

Figure 6.3 Mechanistic detail of the G - dependent mechanism — the cleavage
phosphotransesterifi cation. (Adapted with permission from Figure 1b of reference 16.
Copyright 2003, with permission from Elsevier.)

O

PO

O

B

OH

O 3 ' 3 '

O-

A:H

O OH

O

HO G

B:H

5'

2'

3' δ−

O

P O

O

B

OH

O

O-

O OH

O

HO G

A:H

B:H

5'

2'

3'

δ−

O

OH

B

OH

A:

B:H O OH

O

HO

P O

G

O 3 '

O-

5'

2'

3'