Each primer introduced a terminal NheI site.
Next, the C6″ primary hydroxyl of 1 was selectively reacted with succinic anhydride to introduce a terminal carboxyl function (compound 2) allowing the coupling with the N-terminal group of the fully protected and resin-anchored peptide (compound 3).
In a previous work, we developed a clickable boronic acid, 3- 2-azido-acetylamino phenylboronic acid (APBA) by introducing a terminal azide into commercially available 3- 2-azido-acetylamino phenylboronic
Recombinant expression in Escherchia coli BL21 (DE3) cells was performed from a pET21b vector, which introduces a C-terminal polyhistidine tag.
The vector introduces an amino-terminal myc-epitope tag (EQKLISEEDL) onto the exogenously expressed protein.
This vector introduces an N-terminal His6 tag followed by a TEV protease recognition site (Table S1 of the Supporting Information).
A truncated form lacking the flexible loop spanning amino acids 27 82 and the last 24 amino acids, which are the transmembrane domain, was inserted into the pET15b vector which introduces an N-terminal histidine tag and a thrombin digestion sequence.
The fluorescent mTurquoise2 (donor) and mCitrine (acceptor) proteins were fused to the N-terminal of Kv7.2-ABCD (residues 310-653), and cloned into a pProEX-HTc plasmid (Invitrogen) that introduces a 6xHis N-terminal tag.
Thus, it is reasonable to expect that the effects on the kinetic rates caused by introducing a primer terminal ribose in D12A/D66A complexes will reflect the effects of introducing a primer terminal ribose in wild-type complexes.
The plasmid with ampicillin resistance was amplified with two primers (#1470 and #1471) in order to introduce a C-terminal His6- tag and to delete the C-terminal tryptophan residue of β-lactamase (called ΔW290).
Furthermore, that chapter introduces a heretofore unobserved N-terminal conformation of the group II chaperonin and demonstrates that both the N and C-termini influence the nucleotide usage of the enzyme.
