ACPC deaminase

We noted earlier in this chapter (Section II.C.3) the pyridoxal-phosphate-linked enzymatic route of synthesis of ACPC (9) from the a-aminobutyryl moiety of S-adenosylmethionine (99). ACPC (9) undergoes further metabolic processing by two distinct fragmentation routes, to a-ketobutyrate (138) and ammonia (equation 21) in bacteria and yeast or to ethylene (139) in fruit and other plant tissues (equation 22) where ethylene (139) is a potent hormone for fruit-ripening or wound-healing The bacterial enzyme, ACPC deaminase is much better characterized and is taken up first. 

A detailed set of stereochemical studies have been conducted on the ACPC deaminase reaction. The regiochemistry of ACPC (9) C-C fragmentation was established to be exclusively between the a-C and the Pro-5 P-CR2 group by synthesis and incubation of R-(-)-[2,2-D2]- (9b) and (5)-(-f )-[2,2-D2]-ACPC (9a), which yield [3-D J-(138b) and [4-D2]-2-keto butyrate (138c) respectively Additional stereochemical studies employ... 

To determine the stereoselectivity of diastereotopic proton abstraction from the Pro-R methylene group of ACPC (9) in the fragmentation (occurring between Pro-S p-C and a-C), 2-ethyl-[3-Di]-ACPC (8a) was prepared with the ethyl side chain and deuterium substituent in cis relationship. Incubation of this compound followed by in situ reductive enzymatic trapping with (25)-lactate dehydrogenase yielded 2-hydroxy-[3-D]-hexanoate where the R,5-placement of D was analyzed by NMR and the D-content by mass spectrometry. These results had defined the stereoselectivity for )5-H-abstraction from the Pro-R methylene of 2-ethyl-ACPC (8) as the proton removal in the overall fragmentation process and by analogy the same in ACPC (9). These results place stereochemical constraints on the ACPC deaminase process and were accommodated in Scheme 10. 

A major remaining issue in ACPC deaminase catalysis is the mode of actual cyclopropane fragmentation. At least three mechanistic variants can be pictured (A) p-proton abstraction to initiate cyclopropane fragmentation (B) nucleophilic addition to... 

The ACPC deaminase (EC 4.1.99.4) converts ACPC to 2-oxobutyrate 124, and incubation with (IS)- and (lR)-[2,2- H2]ACPC, which leads to labeling in the methyl and the C-3 positions, respectively, showed that it was the bond to the 2-pro-S carbon that was cleaved in the process (341). This is the bond made on biosynthesis of ACPC. A mechanism for the process is suggested in Scheme 90, and the indication of internal proton return suggests operation... 

The deaminase has been purified by Honma and Shimomura to homogeneity from Pseudomonas and has also been demonstrated to be a PLP-enzyme in some analogy to ACPC synthase. The for ACPC (9) is 9.2 mM and the is 390 min . The cleavage of... 

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