A putative 65-kD PtdInsP kinase protein decreases
in the plasma membrane during the stationary phase. Antisera were
produced in rabbits against a synthetic 15-amino acid peptide
representing a conserved PtdInsP 5 kinase domain from Arabidopsis. A,
Equal protein amounts (20 μg) of microsomes from G.
sulphuraria were separated by SDS-PAGE and blotted to PVDF
membranes. Immunodetection with rabbit pre-immune serum, and with
anti-PtdInsP kinase antiserum, as indicated. Soluble protein extracts
from E. coli expressing AtPIP5KI were separated by SDS-PAGE
and blotted to PVDF membranes. Immunodetection with anti-PtdInsP kinase
antiserum with extract from uninduced cells, or from cells induced for
2.5 h with 1 mm IPTG, as indicated. Blots
stained with amido black are shown (lower). B, Plasma membranes and
microsomes were prepared from G. sulphuraria samples
harvested over the period of time following the transition from
logarithmic growth to the stationary phase. Equal amounts of protein
(20 μg) were separated by SDS-PAGE and blotted to PVDF membranes.
Upper, Immunodetection of PtdInsP kinase protein with the anti-PtdInsP
kinase antiserum. Lower, Blot stained with amido black. White arrows
indicate Mr markers.