L-Lysine → Salicylic Acid

To characterize MS59 further its amino acid sequence was partially determined. Therefore, MS59 was separated in the presence of 0.1 mM thioglycolate in the upper reservoir buffer and SDS on a 12.5% polyacrylamide gel, which was prerun for 2 hours at 50 V with 0.05 mM glutathione in the upper reservoir buffer. The gel was stained with 5% (w/v) Serva Blue G in 45% (v/v) methanol and 10% acetic acid for 30 minutes and destained in 20% (v/v) acetic acid for 30 minutes and the 59 kDa band was cut out and sequenced using Edman degradation on an Applied Biosystems 477A protein sequencer according to the protocol provided by the manufacturer. N-terminal amino acid sequencing of MS59 revealed that the N-terminus was blocked. To obtain internal sequences, MS59 was digested with trypsin. Trypsin cleaves protein at arginine and lysine residues. The digestion products were separated on a reversed-phase column and analyzed by Edman degradation. Two tryptic fragments were sequenced: Pep1 and Pep2. Of Pep1 25 amino acid residues were identified: S-I-N-V-D-I-E-Q-E-T-A-W-V-Q-A-G-A-T-L-G-E-V-Y-Y-R (SEQ ID NO: 1).

1. customTherefore, MS59 was separated in the presence of 0.1 mM thioglycolate in the upper reservoir buffer
2. waitThe gel was stained with 5% (w/v) Serva Blue G in 45% (v/v) methanol and 10% acetic acid for 30 minutes
3. waitdestained in 20% (v/v) acetic acid for 30 minutes
4. customprovided by the manufacturer
5. customTo obtain internal sequences, MS59
6. customThe digestion products were separated on a reversed-phase column

ORD record

ord-b09f75988be7495a82042fd0536aad5e

Source dataset

uspto-grants-2005_03

DOI

10.6084/m9.figshare.5104873.v1

Patent number

US06864076B2