INTERMEDIATES IN THE PHOTOCONVERSION OF FUNCTIONAL PHYTOCHROME IN FERN SPORES OF Dryopteris–II. In vivo KINETICS OF THE DECAY OF Iⁱ₇₀₀ AND OF THE FORMATION OF P_fr STUDIED WITH A DOUBLE‐LASER APPARATUS*

Abstract

Abstract— Photoconversion of the red‐light‐absorbing form of phytochrome, Pr, to the far‐red‐light‐absorbing form, Pfr, was investigated in vivo at 22°C with 600 or 800 ns laser pulses of high spectral purity and induction of spore germination in Dryopteris paleacea was used as indicator for the progress of photoconversion. This reaction is initiated by a saturating R‐laser pulse of 648.5 nm, establishing an equilibrium of the photochromic system between Pr and the very early intermediates, Ii700 (Prφ Ii700)‐ The decay of Ii700 as well as the formation of Pfr was recorded by the application of a second pulse varied between 698 and 717.5 nm, which inhibits the formation of Plr being absorbed predominantly by Ii700or Pfr, respectively. The most effective inhibition for the second pulse is found up to 10 u.s after the first pulse and this is interpreted by photoreversion of Ii700 to Pr; thus reducing the formation of Pfr from Ii700. This early inhibition decreases between 10 μs to 100 ms after the R‐laser pulse, as a result of the decay of Iibl to a bleached species I,;. This decay can be described by three first order kinetics with the rate constants k12= 16830 ± 2970 s‐1, k12= 666 ± 218 s‐1,k13= 9.8 ± 0.9 s‐1. A second inhibition, due to the formation of Pfr, is found for dark intervals <100 ms and can be described by two first order kinetics with the rate constants k21= 2.9 ± 0.6 s‐1 and k22= 0.17 s‐l.

A PHYTOCHROME STUDY USING TWO‐LASER/TWO‐COLOR FLASH PHOTOLYSIS: I700 IS A MANDATORY INTERMEDIATE IN THE PrPfr PHOTOTRANSFORMATION

“…2B trace b) indicating that L2 drives I,oo back to P, as already observed in vitro9 and in vivo.8. 21 From the results of this particular study, we cannot make any assessment about the detailed mechanism of the I,,, decay. The interpretation of our present two-color/two-flash data is independent of the model used to interpret the multiexponential decay of I, , , .…”