Abstract
The pncB gene of Salmonella typhimurium, encoding nicotinate phosphoribosyltransferase (NAPRTase), was cloned on a 4.7-kb Sau3A fragment. The gene contains a 1,200-bp open reading frame coding for a 400-residue protein. Amino acid sequencing of the amino-terminal and two interior peptides of the purified protein confirmed the deduced sequence and revealed that the amino-terminal methionine residue was removed, giving a 399-residue mature protein of Mr 45,512. No signal sequence was observed in the predicted NAPRTase primary structure, suggesting that the enzyme is not periplasmic. The protein does not demonstrate clear sequence similarity to the other seven phosphoribosyltransferases of known primary structure and frustrates attempts to define a consensus 5-phosphoribosyl-1-pyrophosphate-binding region. The NAPRTase reaction is ATP stimulated, and the protein contains a carboxy-terminal sequence diagnostic of an ATP-binding site. An inverted repeat of the sequence TAAACAA observed in the proposed promoter region of pncB is also present in the promoter of nadA, which, like pncB, is also regulated by the NadR (NadI) repressor. The sequence may thus define an NadR repressor-binding site.
Highlights
Cloning and Nucleic Acid Sequence of the Salmonella typhimurium pncB Gene and Structure of Nicotinate Phosphoribosyltransferase
The pncB gene of Escherichia coli and Salmonella typhimurium encodes the enzyme nicotinate phosphoribosyltransferase (NAPRTase; EC 2.4.2.11), which catalyzes the formation of nicotinate mononucleotide (NAMN) [18, 31]
The NAPRTase sequence has been examined for similarity to nucleotide-binding proteins
Summary
Cloning and Nucleic Acid Sequence of the Salmonella typhimurium pncB Gene and Structure of Nicotinate Phosphoribosyltransferase. The protein does not demonstrate clear sequence similarity to the other seven phosphoribosyltransferases of known primary structure and frustrates attempts to define a consensus 5-phosphoribosyl-1-pyrophosphate-binding region. The pncB gene of Escherichia coli and Salmonella typhimurium encodes the enzyme nicotinate phosphoribosyltransferase (NAPRTase; EC 2.4.2.11), which catalyzes the formation of nicotinate mononucleotide (NAMN) [18, 31]. The pncB locus maps at 25 min on the S. typhimurium chromosome [7, 9] and is under the control of a repressor, the product of the nadR gene (8; called nadl [32]), which serves to regulate the expression of the nadA and nadB genes. Our own studies on the S. typhimurium NAPRTase have shown that it catalyzes a slow NAMN synthesis reaction in the absence of ATP. When ATP is present, it is hydrolyzed, and the ATP phosphohydrolase reaction increases the equilibrium constant for NAMN synthesis by about 4,000-fold (unpublished data)
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