Part:BBa_J45001

SAM:salicylic acid carboxyl methyltransferase; converts salicylic acid to methyl salicylate (winter

From snapdragon (A. Majus)

(GenBank Accession No. AF515284)
It is a homodimer with subunit molecular mass of about 45 kDA
On the pET-28a expression vector there is a convenient NdeI site at the initiating ATG codon as well as a BamHI site downstream of the stop codon. Natalia Dudareva is sending the pET-28a vector
concentration: The culture medium of the E. coli cells expressing this SAMT contained methyl salicylate (2.1 μg/ml) when the growing medium was supplemented with 5 μg/ml salicylic acid. Also, they contained methyl benzoate (0.86 μg/ml) when the growing medium was supplemented with 5 μg/ml benzoic acid
pH optimum from 7.0 to 7.5, but the expressed recombinant SAMT enzyme still has 80%-90& maximum activity in pH from 5.0 to 8.0. The enzyme was active in both Tris- and phosphate-citrate-based buffers, although phosphate was 20% lower. Incubation for 30 mins at 42 C had no effect on activity, showing that this SAMT has the broadest temperature stability of this family of enzymes. Also, SAMT activity was not affected by the presence of 5mM Mg2+, K+, NH4+, or Ca2+ in assay reaction.
It does not require any metal ions as cofactors
Fe2+ and Cu2+ have a strong inhibitory effect
Because we are receiving the gene for this enzyme on a pET-28a plamid, the gene is transcribed by T7 polymerase rather than by E. coli polymerase. T7 polymerase is much faster than E. coli so we can expect to get very high expression levels.

Sequence and Features

Assembly Compatibility:

10
COMPATIBLE WITH RFC[10]
12
INCOMPATIBLE WITH RFC[12]
Illegal NheI site found at 577
21
INCOMPATIBLE WITH RFC[21]
Illegal XhoI site found at 901
23
COMPATIBLE WITH RFC[23]
25
INCOMPATIBLE WITH RFC[25]
Illegal NgoMIV site found at 421
Illegal NgoMIV site found at 425
1000
INCOMPATIBLE WITH RFC[1000]
Illegal SapI site found at 471
Illegal SapI.rc site found at 75