TY - JOUR
T1 - Cloning, functional expression, and complementation analysis of an inorganic pyrophosphatase from Bartonella bacilliformis
AU - Mitchell, Samuel J.
AU - Minnick, Michael F.
PY - 1997
Y1 - 1997
N2 - We have cloned the inorganic pyrophosphatase gene (ppa) from the facultative intracellular pathogen Bartonella bacilliformis and characterized its encoded product. The 531-bp gene is located approximately 1 kb downstream of, and in opposite orientation to, the invasion-associated locus (ialAB) of B. bacilliformis. The predicted protein encoded by ppa is 177 amino acid residues, which is in agreement with in vitro and in vivo synthesis of a protein with an apparent molecular mass of 22-23 kDa. The predicted B. bacilliformis pyrophosphatase (PPase) sequence is 53% identical and 85% similar to the E. coli PPase (EC 3.6.1.1), and contains all 12 of the amino acid residues implicated in the catalytic active site. The isolated B. bacilliformis PPase exhibits an activity of 51 ± 2 μmol PO4 released/(mg protein ̇ min) at 28°C and pH 8, and is sensitive to inhibition by Ca2+. In keeping with other prokaryotic PPases, B. bacilliformis PPase activity occurs from pH 6 to 10 (optimal pH = 8) and demonstrates high thermostability in the presence of Mg2+ (highest activity at 55°C, relative activity = 80 ± 3% at pH 8). The cloned B. bacilliformis ppa is able to genetically complement a ppa- mutant strain of E. coli.
AB - We have cloned the inorganic pyrophosphatase gene (ppa) from the facultative intracellular pathogen Bartonella bacilliformis and characterized its encoded product. The 531-bp gene is located approximately 1 kb downstream of, and in opposite orientation to, the invasion-associated locus (ialAB) of B. bacilliformis. The predicted protein encoded by ppa is 177 amino acid residues, which is in agreement with in vitro and in vivo synthesis of a protein with an apparent molecular mass of 22-23 kDa. The predicted B. bacilliformis pyrophosphatase (PPase) sequence is 53% identical and 85% similar to the E. coli PPase (EC 3.6.1.1), and contains all 12 of the amino acid residues implicated in the catalytic active site. The isolated B. bacilliformis PPase exhibits an activity of 51 ± 2 μmol PO4 released/(mg protein ̇ min) at 28°C and pH 8, and is sensitive to inhibition by Ca2+. In keeping with other prokaryotic PPases, B. bacilliformis PPase activity occurs from pH 6 to 10 (optimal pH = 8) and demonstrates high thermostability in the presence of Mg2+ (highest activity at 55°C, relative activity = 80 ± 3% at pH 8). The cloned B. bacilliformis ppa is able to genetically complement a ppa- mutant strain of E. coli.
KW - Bartonella
KW - Inorganic pyrophosphatase
KW - Invasion-associated locus
UR - http://www.scopus.com/inward/record.url?scp=0030819571&partnerID=8YFLogxK
U2 - 10.1139/m97-106
DO - 10.1139/m97-106
M3 - Article
C2 - 9304784
AN - SCOPUS:0030819571
SN - 0008-4166
VL - 43
SP - 734
EP - 743
JO - Canadian Journal of Microbiology
JF - Canadian Journal of Microbiology
IS - 8
ER -