Abstract
Transglutaminase 2 (TG2)
is a distinctive member of the family of Ca2 -dependent enzymes recognized
mostly by their abilities to catalyze the posttranslational crosslinking of
proteins. TG2 uniquely binds and hydrolyzes GTP; binding GTP inhibits its crosslinking
activity but allows it to function in signal transduction (hence the Gh designation).
The core domain of TG2 (residues 139–471, rat) comprises the papain-like
catalytic triad and the GTP-binding domain (residues 159–173) and contains
almost all of the conserved tryptophans of the protein. Examining point
mutations at Trp positions 180, 241, 278, 332, and 337 showed that, upon binding
2 -(or 3 )-O-(N-methylanthraniloyl)GTP (mantGTP), the Phe-332
mutant was the weakest (35% less than wild type) in resonance energy transfer
from the protein ( exc, max 290
nm) to the mant fluorophore ( em 444 nm) and had a reduced affinity for mantGTP.
Trp-332, situated near the catalytic center and the nucleotide-binding area of
TG2, may be part of the allosteric relay machinery that transmits negative
effector signals from nucleotide binding to the active center of TG2. A most
important observation was that, whereas no enzyme activity could be detected
when Trp-241 was replaced with Ala or Gln, partial preservation of catalytic
activity was seen with substitutions by Tyr > Phe > His. The results
indicate that Trp-241 is essential for catalysis, possibly by stabilizing the
transition states by H-bonding, quadrupole–ion, or van der Waals interactions.
This contrasts with the evolutionarily related papain family of cysteine
proteases, which uses Gln-19 (papain) for stabilizing the transition state.
Comment
This paper from Laszlo Lorand's laboratory at Northwestern University, in
collaboration with the lab of Robert Graham, investigated the possible
functional significance of a tryptophan residue in the active site of
transglutaminase. The active
site chemistry appears to be based on a thiol catalytic triad, but the
tryptophan residue may provide the anion hole that stabilizes the transition
state of nucleophilic attack.