Method for isolating mutants overproducing nicotinamide adenine dinucleotide and its precursors.
A procedure has been developed for isolating mutants which are defective with respect to nicotinamide adenine dinucleotide (NAD) metabolism. It is based on the well known V-factor requirement of Haemophilus parainfluenzae. This procedure was used to isolate a series of mutants from Escherichia coli. The pyridine metabolism of wild-type and mutant E. coli cells falls in one of four distinct classes. Class A includes wild-type E. coli and represents strains that are normal with respect to pyridine metabolism. Class B mutants have altered internal pools of NAD. The intracellular NAD concentration of different class B mutants varies over a 10-fold range. Class C mutants excrete pyridine mononucleotides, and class D mutants excrete NAD. The production of pyridine nucleotides by class C and D mutants exceeds that of wild-type E. coli by a factor of at least ten. The mutant strains generally have normal generation times and achieve normal cell densities in minimal medium.
D001431:Bacteriological Techniques; D002247:Carbon Isotopes; D002474:Cell-Free System; D002845:Chromatography; D003470:Culture Media; D004586:Electrophoresis; D004926:Escherichia coli / Q000145:classification / Q000254:growth & development / Q000302:isolation & purification* / Q000378:metabolism*; D005374:Filtration / Q000295:instrumentation; D005827:Genetics, Microbial; D006190:Haemophilus / Q000254:growth & development / Q000378:metabolism; D008722:Methods; D009153:Mutagens; D009154:Mutation; D009243:NAD / Q000096:biosynthesis*; D009536:Niacinamide / Q000378:metabolism; D009539:Nicotinic Acids / Q000378:metabolism; D009604:Nitrosoguanidines; D011742:Pyrimidine Nucleotides / Q000096:biosynthesis; D013696:Temperature
D002247:Carbon Isotopes; D003470:Culture Media; D009153:Mutagens; D009539:Nicotinic Acids; D009604:Nitrosoguanidines; D011742:Pyrimidine Nucleotides; D009243:NAD; D009536:Niacinamide