Editing Aspartate transaminase (section)

==Mechanism==
Aspartate transaminase, as with all transaminases, operates via dual substrate recognition; that is, it is able to recognize and selectively bind two amino acids (Asp and Glu) with different side-chains.<ref name="pmid15889412">{{Cite journal |vauthors=Hirotsu K, Goto M, Okamoto A, Miyahara I |year=2005 |title=Dual substrate recognition of aminotransferases |journal=Chemical Record |volume=5 |issue=3 |pages=160–172 |doi=10.1002/tcr.20042 |pmid=15889412}}</ref> In either case, the transaminase reaction consists of two similar half-reactions that constitute what is referred to as a [[ping-pong mechanism]].  In the first half-reaction, amino acid 1 (e.g., L-Asp) reacts with the enzyme-PLP complex to generate ketoacid 1 (oxaloacetate) and the modified enzyme-PMP.  In the second half-reaction, ketoacid 2 (α-ketoglutarate) reacts with enzyme-PMP to produce amino acid 2 (L-Glu), regenerating the original enzyme-PLP in the process.  Formation of a racemic product (D-Glu) is very rare.<ref name="pmid1735441">{{Cite journal |vauthors=Kochhar S, Christen P |date=February 1992 |title=Mechanism of racemization of amino acids by aspartate aminotransferase |journal=European Journal of Biochemistry |volume=203 |issue=3 |pages=563–569 |doi=10.1111/j.1432-1033.1992.tb16584.x |pmid=1735441 |doi-access=free}}</ref>

The specific steps for the half-reaction of Enzyme-PLP + aspartate ⇌ Enzyme-PMP + oxaloacetate are as follows (see figure); the other half-reaction (not shown) proceeds in the reverse manner, with α-ketoglutarate as the substrate.<ref name="pmid6143829" /><ref name="Biochemistry" />

[[File:Aspartate aminotransferase mechanism.png|class=skin-invert-image|thumb|center|1000px|Reaction mechanism for aspartate aminotransferase]]

# Internal [[aldimine]] formation:  First, the ε-amino group of Lys258 forms a [[Schiff base]] linkage with the aldehyde carbon to generate an internal aldimine.
# Transaldimination:  The internal aldimine then becomes an external aldimine when the ε-amino group of Lys258 is displaced by the amino group of aspartate.  This transaldimination reaction occurs via a [[nucleophilic attack]] by the deprotonated amino group of Asp and proceeds through a tetrahedral intermediate.  As this point, the carboxylate groups of Asp are stabilized by the [[guanidinium]] groups of the enzyme's Arg386 and Arg 292 residues.
# [[Quinonoid zwitterion|Quinonoid]] formation:  The hydrogen attached to the a-carbon of Asp is then abstracted (Lys258 is thought to be the proton acceptor) to form a quinonoid intermediate.
# [[Ketimine]] formation:  The quinonoid is reprotonated, but now at the aldehyde carbon, to form the ketimine intermediate.
# Ketimine [[hydrolysis]]:  Finally, the ketimine is hydrolyzed to form PMP and oxaloacetate.

This mechanism is thought to have multiple partially [[rate-determining step]]s.<ref name="pmid8611515">{{Cite journal |vauthors=Goldberg JM, Kirsch JF |date=April 1996 |title=The reaction catalyzed by Escherichia coli aspartate aminotransferase has multiple partially rate-determining steps, while that catalyzed by the Y225F mutant is dominated by ketimine hydrolysis |journal=Biochemistry |volume=35 |issue=16 |pages=5280–5291 |doi=10.1021/bi952138d |pmid=8611515}}</ref>  However, it has been shown that the substrate binding step (transaldimination) drives the catalytic reaction forward.<ref name="pmid12488449">{{Cite journal |vauthors=Hayashi H, Mizuguchi H, Miyahara I, Nakajima Y, Hirotsu K, Kagamiyama H |date=March 2003 |title=Conformational change in aspartate aminotransferase on substrate binding induces strain in the catalytic group and enhances catalysis |journal=The Journal of Biological Chemistry |volume=278 |issue=11 |pages=9481–9488 |doi=10.1074/jbc.M209235200 |pmid=12488449 |doi-access=free}}</ref>