TABLE 3.
Comparison of arginine cluster sequences in arginine-deficient L. plantarum natural auxotrophs and in arginine prototroph spontaneous derivativesa
| Protein | No. of amino acids | Major mutations in strain(s)b:
|
Minor mutations in strain(s)c:
|
||||||
|---|---|---|---|---|---|---|---|---|---|
| NCFB 2171, NCFB 965, and NCFB 963 | KOG5 | NCFB 772 | R-NCFB772 | FB400, R-FB400-D7, and R-FB400-B4 | R-FB400-A1 | CCM 3626 | R-CCM3626-A | ||
| ArgC | 341 | M14-P341del (deletion; 4915-4997)d | A234V (GCA → GTA; 5573) | A234V (GCA → GTA; 5573), G333E (GGA → GAA; 5870)e | A234V (GCA → GTA; 5573), E333G (GAA → GGA; 5870)f | V35M (GTG → ATG; 4976) | V35M (GTG → ATG; 4976) | NSg | NS |
| ArgJ | 404 | A179V (GCC → GTC; 6458), A354V (GCC → GTC; 6983) | F16C (TTT → TGT; 5969), G305D (GGC → GAC; 6836) | Y150H (TAC → CAC; 6371) | Y150H (TAC → CAC; 6371) | T143M (ACG → ATG; 6351) | T143M (ACG → ATG; 6351) | NS | NS |
| ArgB | 248 | Q32H (CAG → CAT; 7247) | Q32H (CAG → CAT; 7247), L86P (CTT → CGT; 7409) | Q32H (CAG → CAT; 7247), T50N (ACT → AAT; 7301), A81V (GCG → GTG; 7394) | Q32H (CAG → CAT; 7247), T50N (ACT → AAT; 7301), A81V (GCG → GTG; 7394) | Q51stop (CAG → TAG; 7304)h | stop51Y (TAG → TAT; 7304)i | NS | NS |
| ArgD | 389 | K296E (AAA → GAA; 8782) | 47insY (ins TAT; 8037), K296E (AAA → GAA; 8782), S301A (TCC → GCC; 8797), I330M (ATC → ATG; 8884)j | H62Y (CAT → TAT; 8080), K296E (AAA → GAA; 8782), P364S (CCG → TCG; 8986) | NS | N136T (AAT → ACT; 8302) | N136T (AAT → ACT; 8302) | NS | NS |
| ArgF | 340 | NS | NS | NS | NS | NS | NS | P18S (CCA → TCA; 9110), T257A (ACG → GCG; 9827), Q290stop (CAG → TAG; 9926)k | P18S (CCA → TCA; 9110), T257A (ACG → GCG; 9827), stop290L (TAG → TTG; 9926)l |
The argCJBDF reference sequence used was the sequence of the arginine prototroph strain CCM 1904 (accession number X99978). Nucleic acid differences leading to amino acid changes are underlined. Silent mutations are not shown.
No spontaneous arginine prototrophic derivatives were obtained.
Spontaneous arginine prototrophic revertants were selected. Each revertant was designated by using R- followed by a modified version of the designation of the parental arginine-auxotrophic strain.
The amino acids between M14 and P341 were deleted due to an 82-nucleotide deletion. The mutation led to a truncated protein and therefore to arginine auxotrophy.
The G333E mutation led to a nonfunctional protein and therefore to arginine auxotrophy.
The E333G mutation restored arginine prototrophy.
NS, not sequenced.
The amber mutation led to a nonfunctional protein and therefore to arginine auxotrophy.
The mutation restored arginine prototrophy. Another revertant, R-FB400-B3, was analyzed and had a stop51Q mutation (TAG → CAT) in argB, generating an ArgB protein identical to CCM 1904 ArgB.
ins, insertion. The 47insY mutation led to a nonfunctional protein and therefore to arginine auxotrophy.
The amber mutation led to a nonfunctional protein and therefore to arginine auxotrophy.
The stop290L mutation led to a functional protein and therefore to arginine prototrophy. Two other revertants were analyzed: R-CCM3626-K (stop290E; TAG → CAG) and R-CCM3626-P (stop290Y; TAG → TAT).