Integrins
![](/images/gifchars/alpha.gif)
9
![](/images/gifchars/beta2.gif)
1 and
![](/images/gifchars/alpha.gif)
4
![](/images/gifchars/beta2.gif)
1 form a distinct structural class, but while
![](/images/gifchars/alpha.gif)
4
![](/images/gifchars/beta2.gif)
1 has been subjectedto extensive study,
![](/images/gifchars/alpha.gif)
9
![](/images/gifchars/beta2.gif)
1 remains poorly characterized. We have used the small molecule
N-(benzenesulfonyl)-(
L)-prolyl-(
L)-
O-(1-pyrrolidinylcarbonyl)tyrosine (
3) to investigate the biochemical propertiesof
![](/images/gifchars/alpha.gif)
9
![](/images/gifchars/beta2.gif)
1 and directly compare these properties with those of
![](/images/gifchars/alpha.gif)
4
![](/images/gifchars/beta2.gif)
1. Compound
3 has a high affinity forboth integrins with
KD values of
![](/images/entities/le.gif)
3 and 180 pM for
![](/images/gifchars/alpha.gif)
9
![](/images/gifchars/beta2.gif)
1 in 1 mM Mn
2+(activating) and 1 mM Ca
2+and 1 mM Mg
2+ (nonactivating) conditions and
![](/images/entities/le.gif)
5 and 730 pM for
![](/images/gifchars/alpha.gif)
4
![](/images/gifchars/beta2.gif)
1 under the correspondingconditions. Ca
2+ treatment promoted the binding of
3 to both integrins (EC
50 = 30
![](/images/entities/mgr.gif)
M Ca
2+ in bothcases). Compound
3 binding to both integrins was also stimulated by the addition of the activatingmonoclonal antibody TS2/16. These findings indicate that the mechanisms by which metal ions andTS2/16 regulate ligand binding to
![](/images/gifchars/alpha.gif)
9
![](/images/gifchars/beta2.gif)
1 and
![](/images/gifchars/alpha.gif)
4
![](/images/gifchars/beta2.gif)
1 are similar. The binding of
3 to both integrins inducedthe mAb 9EG7 LIBS epitope, a property consistent with occupancy of the receptor's ligand binding siteby
3. But whereas EGTA treatment inhibited the binding of 9EG7 to
![](/images/gifchars/alpha.gif)
4
![](/images/gifchars/beta2.gif)
1, it stimulated the binding of9EG7 to
![](/images/gifchars/alpha.gif)
9
![](/images/gifchars/beta2.gif)
1. The 9EG7 and TS2/16 effects point to contributions of the
![](/images/gifchars/beta2.gif)
1-chains on binding. Cross-linking data revealed that the integrin
![](/images/gifchars/alpha.gif)
-chains are also involved in binding the small molecule, as stablelinkages were observed on both the
![](/images/gifchars/alpha.gif)
9 chain of
![](/images/gifchars/alpha.gif)
9
![](/images/gifchars/beta2.gif)
1 and the
![](/images/gifchars/alpha.gif)
4 chain of
![](/images/gifchars/alpha.gif)
4
![](/images/gifchars/beta2.gif)
1. Extensive structure-activity analyses with natural and synthetic ligands indicate distinct features of the ligand binding pockets.Most notable was the estimated >1000-fold difference in the affinity of the integrins for VCAM-1, whichbinds
![](/images/gifchars/alpha.gif)
4
![](/images/gifchars/beta2.gif)
1with an apparent
KD of 10 nM and
![](/images/gifchars/alpha.gif)
9
![](/images/gifchars/beta2.gif)
1 with an apparent
KD of >10
![](/images/entities/mgr.gif)
M. Differences werealso seen in the binding of
![](/images/gifchars/alpha.gif)
9
![](/images/gifchars/beta2.gif)
1 and
![](/images/gifchars/alpha.gif)
4
![](/images/gifchars/beta2.gif)
1 to osteopontin. Compound
3 competed effectively for thebinding of VCAM-1 and osteopontin to both integrins. While these studies show many similarities in thebiochemical properties of
![](/images/gifchars/alpha.gif)
9
![](/images/gifchars/beta2.gif)
1 and
![](/images/gifchars/alpha.gif)
4
![](/images/gifchars/beta2.gif)
1, they identify important differences in their structure and functionthat can be exploited in the design of selective
![](/images/gifchars/alpha.gif)
9
![](/images/gifchars/beta2.gif)
1 and
![](/images/gifchars/alpha.gif)
4
![](/images/gifchars/beta2.gif)
1 inhibitors.