Please note these web pages are part of an assignment for a graduate course in Advanced Biochemistry and Molecular Biology (BCMB8010) at the University of Georgia. Questions should be directed to Crystal Jackson.

MECHANISM

Viewing Structural Aspects and Mechanism

Link to PDB (Protein Database): 1fkn

http://www.rcsb.org/pdb/cgi/explore.cgi?pdbId=1fkn

Link to gene sequence and PDB Summary Page: 1fkn

http://www.biochem.ucl.ac.uk/bsm/pdbsum/1fkn/main.html

Mechanism

The general reaction scheme for hydrolysis is as follows:

While the specific mechanism of catalytic reaction of -secretase has not been describe in detail, it can be assumed that the proposed mechanism would resemble the mechanism for asptartyl proteases which can be viewed at the following link:

http://prolysis.phys.univ-tours.fr/Prolysis/Images/mecaasp.gif

Aspartyl proteases are bilobed with the active site located between the lobes. Each lobe contributes one aspartate residue of the catlaytic diad. The two aspartyl residues are close in proximity and one residue is ionized, while the other is unionized at the optimum acidic pH range-2-3.
Catalysis by aspartic proteinases involves a nucleophilic attack that is completed by two proton transfers. One is from a water molecule to the diad of the two carboxyl groups and the other is from the diad to the carbonyl oxygen of the substrate with CO-NH cleavage. This is general acid-base catalysis, in which a "push-pull" mechanism leads to the formation of a noncovalent neutral tetrahedral intermediate.

 

CHAINS A AND B OF BETA-SECRETASE IN BLUE AND CYAN.

COORDINATING CHAINS C AND D OF THE INHIBITOR IN GREEN AND ORANGE.

 

Inhibitor OM99-2

2 chains, each 8 residues long: GLU-VAL-ASN-LOL-ALQ-ALA-GLU-PHE

LOL=Leucinol:2(C6H15N1O1)

ALQ=2-methyl-propionic acid:2)C4H8O2)

Modified from PDB Summary, 1fkn, schematic diagram http://www.biochem.ucl.ac.uk/bsm/pdbsum/1fkn/lig01.html.

 

CONSERVED ACTIVE SITE RESIDUES OF CHAINS A AND B OF BETA-SECRETASE: ASP228

Active site residues sit in the center of the cleft along with their surrounding H-bond network.

 

DISULFIDE BRIDGES

There are six Cys residues involved in three disulfide pairs: cys155-359, cys319-269, and cys382-217.

This unique display of disulfide bridges makes beta-secretase a prime target for drug therapy in AD patients.

 

 

Carboxyl Extension

Residues 369-376 form a beta domain with 7 hbonds to strand 293-299. Residues 378-383 form a helix.