Nucleic Acids Research, Vol. 18, No. 15 4595
.=) 1990 Oxford University Press
Nucleotide sequence of the Bacillus subtilis argC gene encoding N-acetylglutamate-gamma-semialdehyde dehydrogenase Margaret C.M.Smith*, Andrew Mountain+ and Simon Baumberg Department of Genetics, University of Leeds, Leeds LS2 9JT, UK EMBL accession
Submitted June 29, 1990
The B. subtilis argC gene encoding N-acetylglutamate-gammasemialdehyde dehydrogenase is required for the third step in the biosynthesis of arginine. argC lies within the argCAEBD-cpaargF cluster at approximately 100 degrees on the chromosome and is expressed from an arginine regulated promoter located just upstream of argC (1, 2, 3). The argC gene was originally cloned by complementation of an argC- arginine auxotroph of E. coli with a 1474 bp insert in pBR322 to form pUL710 (1). TnS insertions in pUL710 abolished the complementation by the cloned gene and localised the argC gene on the insert DNA (1). Analysis of plasmid encoded products in E. coli minicells using pUL710 and TnS insertions indicated the synthesis of a 41 kd protein (M.C.M.Smith, A.Mountain, S.Baumberg, unpublished observations). The cloned insert in pUL71O has been completely sequenced using the Sanger chain termination method on both strands of overlapping Sau3A, TaqI and HpaII clones in M13 (4). The sequence presented here represents the only ORF of the predicted size encoded by the insert. The derived amino acid sequence has 39% identity and 62.5% similarity to the E. coli argC sequence (5). The use of the assigned initiation codon, TTG, has been discussed previously (3). The predicted amino acid sequence encoded between 535 and 561 bp (underlined) is completely conserved between the E. coli and the B. subtilis sequences and is similar to 'Box A' of the nucleotide binding pocket consensus sequence (6).
1
TTGAAAATAGGAATTGTAGGTGCTACAGGATATGGAGGCACCGAACTTGTCAGGATTCTT AACTTTTATCCTTAACATCCACGATGTCCTATACCTCCGTGGCTTGAACAGTCCTAAGAA M
61
I
G
I
V
G
A
T
G
Y
G
G
T
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H
A
E
E
C
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S
S
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G
N
V
121
ATATCGCTCCCAATAGGAGTAGAATGGCCGAATCGCCTAGTCGTCGACTTCGGCTAACTA Y S E G Y P H L T G L A D Q Q L K P I D 181
ATGAATACGATCAAACACGAAATAGATATCATGTTTCTCGCTGCGCCGCCCGGAGTATCA TACTTATGCTAGTTTGTGCTTTATCTATAGTACAAAGAGCGACGCGGCGGGCCTCATAGT M
N
T
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K
H
E
I
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I
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A
A
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AGTGAATTGACTCCAAAGCTGGCAGACGCGGGAATTACGGTTATTGATCTGTCAGGTGAT TCACTTAACTGAGGTTTCGACCGTCTGCGCCCTTAATGCCAATAACTAGACAGTCCACTA
301
CTGAGGATAAAAGAACCGGCTGAATATGAAAAATGGTATAAACGGACAGCGGCACCGAAG GACTCCTATTTTCTTGGCCGACTTATACTTTTTACCATATTTGCCTGTCGCCGTGGCTTC
361
GCGGTGATTCAAGAGGCGGTATACGGTCTGGCAGAACTGAATCAACTGCAAATTCAACAG CGCCACTAAGTTCTCCGCCATATGCCAGACCGTCTTGACTTAGTTGACGTTTAAGTTGTC
S L
A
E
R
V
L
I
I
T
K
Q
P E
E
K P
A
L A
V
A
E
Y
D
Y
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A
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G K
A
I
W
E
T
Y
L
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K
N
I
R
Q
D
T
L
L
A
Q
S A
I
G P
Q
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K
Q
421
GCGAAACTCATTGCCAATCCAGGCTGTTTTCCAACAGCTGTTTTGCTTGGCCTCGCGCCA CGCTTTGAGTAACGGTTAGGTCCGACAAAAGGTTGTCGACAAAACGAACCGGAGCGCGGT
481
TTGGCTCAAAAGAAACTGCTCGATGAATCTTTCGTTATCGTTGACGCGAAGACCGGTGTT AACCGAGTTTTCTTTGACGAGCTACTTAGAAAGCAATAGCAACTGCGCTTCTGGCCACAA
541
TCCGGAGCGGGAAGAAAAGCATCCATGGGAACTCATTTTTCTGAGCTGAACGACAATTTT AGGCCTCGCCCTTCTTTTCGTAGGTACCCTTGAGTAAAAAGACTCGACTTGCTGTTAAAA
601
AAAATTTATAAAGTCAATGAACATCAGCACACGCCGGAAATTGAGCAGGCGCTGAATGAA TTTTAAATATTTCAGTTACTTGTAGTCGTGTGCGGCCTTTAACTCGTCCGCGACTTACTT
661
TGGCAGCCAGGTCTCGGGCCCATTACATTTTCGGCTCACTTGTTTCCGATGACAAGGGGC
A
L
S K
W
K
A
G I
Q
L
Q A
Y
P
I
K
G K
G
A
K
R
V
L
N
L
K
N
G
P
L
A
E
P
G D
S H
I
C E
M
Q T
F
S
G H F
P
F
T
T
S
T
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H P
A
A
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F
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H
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E E
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L
Q P
G K
N
A
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T
A
G N
N
R
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V
F
E
G
721
ATCATGGCGACGATGTATACCAGATTAACCTGTGACCTAACCGCAGATGACCTGCATGAT TAGTACCGCTGCTACATATGGTCTAATTGGACACTGGATTGGCGTCTACTGGACGTACTA
781
TTATATTCGGAATTTTACCAAGATTCATATTTTGTGAGAGTGAGGCCAAAAGGTCAGTAC AATATAAGCCTTAAAATGGTTCTAAGTATAAAACACTCTCACTCCGGTTTTCCAGTCATG
841
CCGCAAACGAAAGAAGTGTACGGCAGCAATTTCTGTGATATCGCCGTGACCCTCGATGAG GGCGTTTGCTTTCTTCACATGCCGTCGTTAAAGACACTATAGCGGCACTGGGAGCTACTC
I
P
1. Mann,N.H., Mountain,A., Munton,R.N., Smith,M.C.M. and Baumberg,S. (1983) Mol. Gen. Genet. 197, 75-81. 2. Mountain,A., McChesney,J., Smith,M.C.M. and Baumberg,S. (1986) J. Bacteriol. 165, 1026-1028. 3. Smith,M.C.M., Mountain,A. and Baumberg,S. (1986) Gene 49, 53-60. 4. Messing,J. (1983) Methods Enzymol. 101, 20-78.
K
TCGCATCATCCTCATGCAGAGGAATGCATACTTTATTCATCCAGCGGAGAAGGGAATGTC AGCGTAGTAGGAGTACGTCTCCTTACGTATGAAATAAGTAGGTCGCCTCTTCCCTTACAG
L
REFERENCES
X52834
5. Parsot,C., Byen,A., Cohen,G.N. and Glansdorff,N. (1988) Gene 68, 275-283. 6. Walker,J.E., Saraste,M., Runswick,M.J. and Gay,N.J. (1982) EMBO J. 1, 945-951.
ACKNOWLEDGEMENTS This work was supported by MRC grants G82/0499/8CB to SB and G84/0925/OCB to SB and AM.
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AGAACGAACAGAGTCACGATCGTCTCGGTAATCGATAATTTAATGAAGGGTGCCGCCGGT TCTTGCTTGTCTCAGTGCTAGCAGAGCCATTAGCTATTAAATTACTTCCCACGGCGGCCA R
961
M
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CAGGCAGTGCAAAACTTTAATTTGATGAATGGCTGGAATGAAGAAACTGGACTCACCATC GTCCGTCACGTTTTGAAATTAAACTACTTACCGACCTTACTTCTTTGACCTGAGTGGTAG Q A V Q N F N ACGCCAATTTATCCATAG TGCGGTTAAATAGGTATC T
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*To whom correspondence should be addressed at the Department of Biological and Molecular 'Present address: Celltech Group plc, 216 Bath Road, Slough SLI 4EN, UK
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*
Science, University of Stirling, Stirling FK9 4LA, UK
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