Mj. Butler et al., CLONING AND CHARACTERIZATION OF A GENE ENCODING A SECRETED TRIPEPTIDYL AMINOPEPTIDASE FROM STREPTOMYCES-LIVIDANS-66, Applied and environmental microbiology, 61(8), 1995, pp. 3145-3150
The gene encoding a tripeptidyl aminopeptidase (Tap) from Streptomyces
lividans was cloned by using a simple agar plate activity assay. Over
expression of the cloned gene results in the production of a secreted
protein,which has an apparent subunit molecular weight of 55,000 and i
s responsible for the major aminoterminal degradative activity in cult
ure broths of S. lividans strains, A DNA sequence analysis revealed a
potential protein-encoding region of the size expected to encode the o
bserved protein, which contained a sequence that exhibited significant
homology around a putative active site serine residue observed far li
pases, esterases, and acyl transferases. Preceding the amino terminus
of the secreted protein was a predicted signal peptide of 36 amino aci
ds followed by a tripeptide, which could be autocatalytically removed
from a secreted Tap precursor, The transcriptional start site for the
gene was mapped by primer extension. Mutant strains of S. lividans lac
king detectable Tap activity were able to grow and sporulate normally.
Cross-species hybridization experiments showed that DNA homologs of t
he fap gene are present in most of the Streptomyces strains tested.