Mass spectrometric based mapping of the disulfide bonding patterns of integrin alpha chains.
Integrins are one of the major mediators of cellular adherence. Structurally the component alpha and beta chains are characterized by extensive intrachain disulfide bonding. The assignment of these bonds is currently based on homology with the chains of the integrin alphaIIbbeta3. However, recent crystallographic analysis of the soluble alphaVbeta3 construct indicates that the alphaV chain displays bonding patterns different from those predicted for alphaIIb. In an effort to define the disulfide bonding patterns in integrins, we have used mass spectrometric based approaches to map the human alpha3, alpha5, alphaV, and alphaIIb. The results indicate that there are differences in the disulfide patterns of the alpha chains. These do not correlate with the integrin capacity to bind ligands as all integrins used in the present study displayed functional activity. The differences were observed in the bonding patterns linking the heavy (H) and light (L) components of the of the alpha chains. It was also possible to assign the location in alpha5 of an additional disulfide bond involving a pair of cysteines not present in alphaV or alphaIIb. This second bond between the H and L chains of alpha5 has not been previously described. These results indicate that not all integrin species display the same disulfide bonding patterns. They also highlight the need for caution in the use of assignments based on sequence homology.
Manitoba Centre for Proteomics, University of Manitoba, Winnipeg, Manitoba, Canada R3E 3P4.
Oleg V
OV
Krokhin
Keding
K
Cheng
Sandra L
SL
Sousa
Werner
W
Ens
Kenneth G
KG
Standing
John A
JA
Wilkins
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D003545:Cysteine / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry*; D006801:Humans; D039201:Integrin alpha3beta1 / Q000737:chemistry*; D039081:Integrin alpha5beta1 / Q000737:chemistry*; D039302:Integrin alphaVbeta3 / Q000737:chemistry*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010084:Oxidation-Reduction; D010449:Peptide Mapping / Q000379:methods*; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods
D004220:Disulfides; D039201:Integrin alpha3beta1; D039081:Integrin alpha5beta1; D039302:Integrin alphaVbeta3; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D003545:Cysteine