Role of single disulfide in recombinant human tumor necrosis factor-alpha

Two analogs of tumor necrosis factor-alpha (TNF-alpha) were produced by in vitro site-directed mutagenesis. In these analogs, cysteine residues at positions 69 and 101, which form a disulfide bond, were changed to alanine or leucine. CD spectra showed that the analogs are apparently similar in secon...

Full description

Saved in:
Bibliographic Details
Published in:The Journal of biological chemistry 1987-09, Vol.262 (27), p.13107-13110
Main Authors: Narachi, M A, Davis, J M, Hsu, Y R, Arakawa, T
Format: Article
Language:English
Subjects:
Analytical, structural and metabolic biochemistry
Animals
Biological and medical sciences
Cell Survival - drug effects
Circular Dichroism
Disulfides
Fundamental and applied biological sciences. Psychology
Holoproteins
L Cells - cytology
L Cells - drug effects
Macromolecular Substances
man
Mice
Mutation
Other proteins
Protein Conformation
Proteins
Recombinant Proteins - pharmacology
Spectrometry, Fluorescence
Structure-Activity Relationship
tumor necrosis factor alpha
Tumor Necrosis Factor-alpha - genetics
Tumor Necrosis Factor-alpha - pharmacology
Citations: Items that this one cites
Items that cite this one
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Two analogs of tumor necrosis factor-alpha (TNF-alpha) were produced by in vitro site-directed mutagenesis. In these analogs, cysteine residues at positions 69 and 101, which form a disulfide bond, were changed to alanine or leucine. CD spectra showed that the analogs are apparently similar in secondary and tertiary structure to the natural sequence TNF-alpha. In addition, the molecular size of the analogs was identical to that of the natural sequence TNF-alpha as determined by gel filtration. However, fluorescence spectra and quenching indicated that the removal of the disulfide bond alters the local conformation around tryptophan residues. The cytolytic, macrophage activation, and lipogenic activities decreased in the order of the natural sequence TNF-alpha greater than the alanine analog greater than the leucine analog, suggesting that the surface involving the disulfide bond plays a role in these biological functions and the introduced modifications decrease the activity. Differential effect of the modifications was suggested in the antiviral activity, since in this assay only the leucine analog showed significantly lower activity.
ISSN:0021-9258
1083-351X
DOI:10.1016/S0021-9258(18)45174-5