Efficient N-terminal peptide sequencing using bis(terpyridine)ruthenium(II) derivatives

Rapid and Sensitive Amino-Acid Sequencing of Cloning Thermus thermophilus HB8 Ferredoxin by Proteomics

“…DNTPs and Taq DNA polymerase were obtained from Takara Co. Ampicillin-resistant pT7-blue vecter was taken from Novagen Co. E . coli strain DH5α was purchased from Novagen Co. [(tpy)Ru(tpy-C 6 H 4 −NCS)](PF 6 ) 2 was prepared by the same method described in the previous paper …”

“…DNTPs and Taq DNA polymerase were obtained from Takara Co. Ampicillin-resistant pT7-blue vecter was taken from Novagen Co. E. coli strain DH5R was purchased from Novagen Co. [(tpy)Ru(tpy-C6H4-NCS)](PF6)2 was prepared by the same method described in the previous paper. 13 Cloning and Sequencing of Thermus thermophilus HB8 Ferredoxin (TtFd) Gene. TtFd gene was cloned from Thermus thermophilus HB8 genomic DNA using a polymerase chain reaction (PCR) and two oligonucleotide primers designed according to the ORF of TtFd from the genome.…”

“…To establish the amino acid sequence of holo TtFd obtained by the genetics-based method, a sequence containing four amino acids from N-terminus was determined by an Edman degradation using a novel tpy-Ru(II)-tpy-C 6H4-NCS (〈Ru〉-NCS) Edman reagent 13 that reacts with the Nterminus of SDS-denatured TtFd as shown in Figure 2.…”

Distinction of Leu and Ile Using a Ruthenium(II) Complex by MALDI-LIFT-TOF/TOF-MS Analysis

“…37,38 The reagent has a characteristic isotope pattern and yields highly intense peaks in MS analysis. [37][38][39][40][41] By using this reagent for amino acid sequencing of peptides or proteins, we selectively detected N-terminal fragment ions with no C-terminal fragment ion. N-Terminal labeling with the reagent 〈Ru〉-CO achieves rapid determination of the Nterminal amino acid sequence.…”

“…Recently, we demonstrated protein sequencing using an active ester of bis(terpyridine)ruthenium(II) derivative (Figure ) as a novel N-terminal labeling reagent. , The reagent has a characteristic isotope pattern and yields highly intense peaks in MS analysis. − By using this reagent for amino acid sequencing of peptides or proteins, we selectively detected N-terminal fragment ions with no C-terminal fragment ion. N-Terminal labeling with the reagent 〈Ru 〉 -CO achieves rapid determination of the N-terminal amino acid sequence .…”