Fig 4. Residues at the front entry of the Tgl tunnel are more important than back residues for the localization of the enzyme.
A: Forms of Tgl-CFP bearing the indicated single Ala substitutions (color coded according to their role in activity or position in the structure of Tgl; as in Fig 3) were produced from the amyE locus in a strain bearing a tgl::sp insertional allele. Sporulation was induced by resuspension, samples collected 8 hours thereafter, and the assembly of the WT and different forms of Tgl-CFP monitored by phase contrast and fluorescence microscopy. Scale bar, 1 μm. B: Cells bearing phase bright spores were analyzed and the normalized forespore/cell fluorescence ratio, (FFS/Fcell)norm, was determined (see also Fig 2). For cells of each strain, the individual normalized ratio values, the median, and interquartile range are shown. n, number of sporangia scored. C: The fluorescence signal detected in whole cells was normalized using the median value of the wild type strain grown on the same day, to give (Fcell)norm (see materials and methods for details). The same cells were analyzed in B and C.