Cacao

An534209, IMP is the correct evidence type! Follow the evidence code decision tree: is annotation based on genetic mutation or allelic variation? --Yes!!! (Read the paper before challenging next time!!!)

Incorrect evidence type, it should be IDA (Inferred from direct assay).

Fig. 5, lanes 1-7 are just showing that His does not play a role in the active site of EII in phosphorylation (that's why there was phosphorylation activity). Replacement of His and Cys residues by site-directed mutagenesis resulted in total loss of sugar phosphorylating activity only (key word here) when Cys483 was replaced. It is concluded that Cys483 is the phosphorylated residue in the active center of the EII lac from Lactobacillus casei. This experiment was made through site-directed mutagenesis of the lacE gene to identify that a single amino acid, cysteine, is essential for a lactose phoshorylation. Conclusion: lacE gene is involved in phosphorylation activity. Fig. 5, lanes A-F, lactose was phosphorylated in the presence of complete system (lane C), but not when either membranes (lane B) or PEP (lane D) were omitted from the reaction. Another confirmation that Cys is the essential amino acid in EII for this microorganism is Fig. 7, lanes 10-11: when Cys483 was replaced with His, an amino acid found to be involved in all other phosphoryltransfer reactions of the PTS, or Tyr, no lactose phosphorylation was observed.

Fig. 5 shows that in lanes 1-4, mutant proteins could still transfer the phosphoryl group to lactose, and lanes 5-7 also show that the leucine mutant proteins also are able to transfer the phosphoryl group to lactose. In addition, lanes A-E are the most important lanes because they show that the opposite orientation of transcription driven by Plac were not able to phosphorylate lactose, indicating that lactose phosphorylation was dependent upon the Plac driven transcription of the lacE gene.