Millisecond protein regions

Ffe. 5.11 TLie MM forms of rhodopsin (R) are in equilibrium with their parent compound Ml and the progeny Mill. The equilibnum between the metastates, Ml, Mila, and Mllb, are attained within a few milliseconds after illumination. The Ml/Mlla transition is accompanied by the intramolecular translocation of the Schiff-base proton to GlullS, located in a region close to the lipid membrane. The subsequent Mlla/Mllb transition is dependent on proton uptake from the aqueous phase and occurs in a watersxposed area. Fast binding of the G protein, transducin, occurs only with Mllb.(Reproduced from ref. 58 with permission of Elsevier Science.)... 

Figure 4 A schematic representation of the experimentai approach for time-resoived XAS measurements. XAS provides local structural and electronic information about the nearest coordination environment surrounding the catalytic metal ion within the active site of a metalloprotein in solution. Spectral analysis of the various spectral regions yields complementary electronic and structural information, which allows the determination of the oxidation state of the X-ray absorbing metal atom and precise determination of distances between the absorbing metal atom and the protein atoms that surround it. Time-dependent XAS provides insight into the lifetimes and local atomic structures of metal-protein complexes during enzymatic reactions on millisecond to minute time scales, (a) The drawing describes a conventional stopped-flow machine that is used to rapidly mix the reaction components (e.g., enzyme and substrate) and derive kinetic traces as shown in (b). (b) The enzymatic reaction is studied by pre-steady-state kinetic analysis to dissect out the time frame of individual kinetic phases, (c) The stopped-flow apparatus is equipped with a freeze-quench device. Sample aliquots are collected after mixing and rapidly froze into X-ray sample holders by the freeze-quench device, (d) Frozen samples are subjected to X-ray data collection and analysis.

Fig. 2. (A) Flash-induced AA in the SDS-fractionated PS-1 core complex (CPI) at 5 K [ with and o without DCIP] (B) Flash-induced AA in TSF-I particles containing dithionite and neutral red at pH 10 and frozen while being illuminated (C) left AA induced by 300-ns, dye laser flashes [710 nm for the blue and green region 590 nm for the red region] insets show individual AA transients at 696 and 480 nm (C) right The difference between the difference spectrum in the left panel and that of P700. (D) Plot of the rate constant vs. reciprocal temperature. Figure source (A) Mathis, Sauer and Remy (1978) Rapidly reversible flash-induced electron transfer on a P-700 chlorophyll-protein complex isolated with SDS. FEBS Lett 88 277 (8) Sauer, Mathis, Acker and van Best (1979) Absorption changes of P-700 reversible in milliseconds at low temperature in Triion-solubilized photosystem I particles. Biochim Biophys Acta 545 469 (C and D) Shuvalov, Dolan and Ke (1979) Spectral and kinetic evidence for two eariy electron acceptors in phoiosystem I. Proc Nat Acad Sci, USA 76 771,773.

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