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Assignment of the disulfide bonds in the sweet protein brazzein
Author(s) -
Kohmura Masanori,
Ota Masafumi,
Izawa Hiroyuki,
Ming Hellekant Ding Göran,
Ariyoshi Yasuo
Publication year - 1996
Publication title -
biopolymers
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.556
H-Index - 125
eISSN - 1097-0282
pISSN - 0006-3525
DOI - 10.1002/(sici)1097-0282(199604)38:4<553::aid-bip10>3.0.co;2-b
Subject(s) - chemistry , thermostability , thermolysin , hydrolysis , cystine , substrate (aquarium) , enzyme , enzymatic hydrolysis , amino acid , intramolecular force , disulfide bond , biochemistry , stereochemistry , cysteine , trypsin , oceanography , geology
The thermostable sweet protein brazzein consists of 54 amino acid residues and has four intramolecular disulfide bonds, the location of which is unknown. We found that brazzein resists enzymatic hydrolysis at enzyme/substrate ratios (w/w) of 1:100‐1:10 at 35–40°C for 24–48 h. Brazzein was hydrolyzed using thermolysin at an enzyme/substrate ratio of 1:1 (w/w) in water, pH 5.5. for 6 h and at 50°C. The disulfide bonds were determined, by a combination of mass spectrometric analysis and amino acid sequencing of cystine‐containing peptides, to be between Cys4‐Cys52, Cys16‐Cys37, Cys22‐Cys47, and Cys26‐Cys49. These disulfide bonds contribute to its thermostability. © 1996 John Wiley & Sons, Inc.