Journal: bioRxiv

Article Title: Mechanism of the secretion of the lanthipeptide nisin

doi: 10.1101/839423

Figure Lengend Snippet: ( A ) The supernatants of pre-NisA secreting L. lactis NZ9000 strains was analyzed by RP-HPLC and the amount of pre-NisA was determined. Amounts of secreted pre-peptides (nmol) are plotted against time (min) and the resulting curves were fitted by an allosteric sigmoidal fit. Modified NisA (mNisA, red) was secreted by strain NZ9000BTC (red dots) and can be preclude by nisT deletion (strain NZ9000BC, clear dot) or an ATP-deficient mutant (NZ9000BT H551A C, red rhomb). Dehydrated NisA (dNisA, blue) was secreted by strains NZ9000BTC H331A (blue rhomb) and NZ9000BT (blue dot), whereas unmodified NisA (uNisA, grey) was secreted by the strains NZ9000T (grey dots) and NZ9000TC (grey square). Dashed square shows a zoom-in on strains with lower secretion level. ( B ) The kinetic parameter of V max (nmol) of secreted peptides was plotted as bars against the various secretion systems. ( C ) The secretion rate of NisA molecules per NisT molecules was plotted against time (min) and fitted by linear regression. The slope represented the secretion rate of NisA•NisT −1 •min −1 for the strains NZ9000BTC and NZ9000T. All data represent secretion experiments from at least five different transformants and are represented as means ± SD (n=5). ++: WT secretion; o: low secretion; −: no secretion

Article Snippet: The culture supernatants containing pre-NisA variants were analyzed by RP-HPLC (Agilent Technologies 1260 Infinity II).

Techniques: Modification, Mutagenesis

Journal: bioRxiv

Article Title: Mechanism of the secretion of the lanthipeptide nisin

doi: 10.1101/839423

Figure Lengend Snippet: The supernatants of pre-NisA secreting L. lactis NZ9000 strains were employed for an RP-HPLC analysis. The pre-NisA variants (mNisA, dNisA and uNisA) were separated from other peptides in the supernatant by an acetonitrile/water gradient on a C-18 RP-HPLC column (left panel). The elution fractions (dashed square; middle panel) were further analyzed by MALDI-TOF-MS (right panel) to verify the correct mass. Integration of the corresponding peaks enables the determination of peptide amounts (nmol). Supernatant analysis of L. lactis strains ( A ) NZ9000BTC, ( B ) NZ9000BTC H331A , ( C ) NZ9000BT H551A C, ( D ) NZ9000BT, ( E ) NZ9000T, ( F ) NZ9000TC and ( G ) NZ9000BC.

Article Snippet: The culture supernatants containing pre-NisA variants were analyzed by RP-HPLC (Agilent Technologies 1260 Infinity II).

Techniques: