Functional expression of the signaling complex sensory rhodopsin II/transducer II from Halobacterium salinarum in Escherichia coli.
Sensory rhodopsin II, a photoreceptor from Halobacterium salinarum (HsSRII), in complex with its cognate transducer protein (HsHtrII) triggers the photophobic response via a cytoplasmic two-component signaling cascade. HsHtrII possess in addition to the HsSRII binding and the cytoplasmic domains an extracellular serine-receptor domain. Here we describe the properties of HsSRII and HsHtrII and those of various shortened transducer analogs, heterologously expressed in Escherichia coli. HsSRII displays the photocycle typical of archaeal photosensors with prolonged kinetics. Using an isothermal titration calorimetric analysis for this complex a dissociation constant of 1.1 microm was obtained similar to that of the corresponding transducer/receptor pair from Natronobacterium pharaonis. A shortened transducer lacking the extracellular and cytoplasmic domain is also sufficient to bind the receptor with a slightly lower affinity. The dissociation constant of serine binding to the extracellular domain was determined to be about 5 microm. This result is in line with the proposal that the extracellular domain indeed is a serine receptor.
Kim
Young-Jun
YJ
Max-Planck-Institut für molekulare Physiology, Dortmund, Germany.
Chizhov
Igor
I
Engelhard
Martin
M
D000143:Acids; D002151:Calorimetry; D004926:Escherichia coli / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D019617:Halobacterium salinarum / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D055668:Photochemical Processes; D011485:Protein Binding; D025603:Sensory Rhodopsins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D012694:Serine / Q000737:chemistry; D013816:Thermodynamics; D017186:Titrimetry
D000143:Acids; D025603:Sensory Rhodopsins; D012694:Serine