similarly, Tyr268, Tyr294, Lys51, and Glu295 (Pseudomonas sp. UW4 ACC
deaminase numbering scheme). The possible role of a reactive thiol group of
Cys162, which is located between the two major domains of the molecule, has
also been suggested (Glick 2005). Overall, it has been suggested that Lys51 within
the active site of ACC deaminase is involved in proton extraction from the substrate
(namely ACC), whereas Tyr294 has been proposed as a key catalytic residue that
positions the cofactor PLP to be in the correct orientation within the active site,
helping substrate to bind properly, and facilitating the formation of the external
aldimine between PLP and the substrate by reacting with the amino group of the
substrate (Karthikeyan et al. 2004; Ose et al. 2003).
19.4.1 Optimal Temperature and pH Profile of ACC
Deaminase
The optimal activity of the ACC deaminase enzyme occurs at pH 8–9. Pure enzyme
from bacterium Pseudomonas sp. UW4 exhibited optimal activity at pH 8.0 with a
sharp loss of activity at acidic pH 6.5 and a complete loss of activity at values of pH
above 10, with estimated pK a values of 7.4 and 9.5 (Hontzeas et al. 2004). Similarly,
P. putida GR12-2 ACC deaminase has optimal activity at pH 8.5 and pK a values of
approximately 7.7 and 9.2 (Jacobson et al. 1994) while the ACC deaminase from the
genus Methylobacterium exhibited optimal activity at pH 8.0 (Fedorov et al. 2013).
The enzyme optimal temperature varies hugely among different bacterial genera.
Enzymes from Pseudomonas spp. generally perform optimally at 30
C, with the
exception of strain UW4, which exhibits optimal activity at ~25
C. Nevertheless,
pseudomonad ACC deaminases are thermodynamically stable enzymes (T m ¼ 60
C)
based on their melting curve analysis (Hontzeas et al. 2004). On the other hand, ACC
deaminases from Cyberlindera saturnus, Methylobacterium nodulans ORS2060,
and Methylobacterium radiotolerans JCM2831 were found to be optimally active
at 37
C, 50
C, and 45
C, respectively (Table 19.1).
19.4.2 Effect of Heavy Metals on ACC Deaminase Activity
ACC deaminase-containing bacteria have been documented to promote host plant
growth and development in the presence of heavy metals (Burd et al. 2000) and the
effect of heavy metals on the activity of ACC deaminase has been investigated
(Carlos et al. 2016). In this study, the ACC deaminase activity of ten different
bacterial strains was measured in the presence and absence of heavy metals (Pb,
As, Cu, Ni, Cd, and Mn). It was found that although all of the selected bacteria
contain active ACC deaminase, there were differences in the way that metals
affected those bacteria. For instance, the ACC deaminase activity of bacterial strains
19 Biochemistry and Molecular Biology of the Enzyme ACC Deaminase
373
deaminase numbering scheme). The possible role of a reactive thiol group of
Cys162, which is located between the two major domains of the molecule, has
also been suggested (Glick 2005). Overall, it has been suggested that Lys51 within
the active site of ACC deaminase is involved in proton extraction from the substrate
(namely ACC), whereas Tyr294 has been proposed as a key catalytic residue that
positions the cofactor PLP to be in the correct orientation within the active site,
helping substrate to bind properly, and facilitating the formation of the external
aldimine between PLP and the substrate by reacting with the amino group of the
substrate (Karthikeyan et al. 2004; Ose et al. 2003).
19.4.1 Optimal Temperature and pH Profile of ACC
Deaminase
The optimal activity of the ACC deaminase enzyme occurs at pH 8–9. Pure enzyme
from bacterium Pseudomonas sp. UW4 exhibited optimal activity at pH 8.0 with a
sharp loss of activity at acidic pH 6.5 and a complete loss of activity at values of pH
above 10, with estimated pK a values of 7.4 and 9.5 (Hontzeas et al. 2004). Similarly,
P. putida GR12-2 ACC deaminase has optimal activity at pH 8.5 and pK a values of
approximately 7.7 and 9.2 (Jacobson et al. 1994) while the ACC deaminase from the
genus Methylobacterium exhibited optimal activity at pH 8.0 (Fedorov et al. 2013).
The enzyme optimal temperature varies hugely among different bacterial genera.
Enzymes from Pseudomonas spp. generally perform optimally at 30
C, with the
exception of strain UW4, which exhibits optimal activity at ~25
C. Nevertheless,
pseudomonad ACC deaminases are thermodynamically stable enzymes (T m ¼ 60
C)
based on their melting curve analysis (Hontzeas et al. 2004). On the other hand, ACC
deaminases from Cyberlindera saturnus, Methylobacterium nodulans ORS2060,
and Methylobacterium radiotolerans JCM2831 were found to be optimally active
at 37
C, 50
C, and 45
C, respectively (Table 19.1).
19.4.2 Effect of Heavy Metals on ACC Deaminase Activity
ACC deaminase-containing bacteria have been documented to promote host plant
growth and development in the presence of heavy metals (Burd et al. 2000) and the
effect of heavy metals on the activity of ACC deaminase has been investigated
(Carlos et al. 2016). In this study, the ACC deaminase activity of ten different
bacterial strains was measured in the presence and absence of heavy metals (Pb,
As, Cu, Ni, Cd, and Mn). It was found that although all of the selected bacteria
contain active ACC deaminase, there were differences in the way that metals
affected those bacteria. For instance, the ACC deaminase activity of bacterial strains
19 Biochemistry and Molecular Biology of the Enzyme ACC Deaminase
373