2 research outputs found
Conceptional Design of Self-Assembling Bisubstrate-like Inhibitors of Protein Kinase A Resulting in a Boronic Acid Glutamate Linkage
The spontaneous esterification of
boronic acids with polyols provides
a promising opportunity to generate self-assembled bisubstrate-like
inhibitors within the binding pocket of cAMP-dependent protein kinase
(PKA). As a first substrate component, we designed amino acids, which
have either a boronic acid or ribopyranose side chain and introduced
them to the substrate-like peptide protein kinase inhibitor (PKI).
The second component was derived from the active-site inhibitor Fasudil,
which was functionalized with phenylboronic acid. NMR spectroscopy
in dimethylsulfoxide proved spontaneous reversible condensation of
both components. Reinforced by the protein environment, both separately
bound substrates were expected to react via boronic-ester formation
bridging the two binding sites of PKA. Multiple crystal structures
of PKA with bound PKIs, positionally modified with residues such as
a ribopyranosylated serine and threonine or a phenylboronic acid attached
to lysine via amide bonds, were determined with the phenylboronic
acid-linked Fasudil. Although PKA accepts both inhibitors simultaneously,
the expected covalent attachment between both components was not observed.
Instead, spontaneous reaction of the terminal boronic acid group of
the modified Fasudil with the carboxylate of Glu127 was detected once
the latter residue is set free from a strong salt bridge formed with
arginine by the original peptide inhibitor PKI. Thus, the desired
self-assembly reaction occurs spontaneously in the protein environment
by an unexpected carboxylic acid boronate complex. To succeed with
our planned self-assembly reaction between both substrate components,
we have to redesign the required reaction partners more carefully
to finally yield the desired bisubstrate-like inhibitors in the protein
environment