Ca2+-Triggering

Fig. 4.1.3 Absorption spectra of aequorin (A), spent solution of aequorin after Ca2+-triggered luminescence (B), and the chromophore of aequorin (C). Fluorescence emission spectrum of the spent solution of aequorin after Ca2+-triggered bioluminescence, excited at 340 nm (D). Luminescence spectrum of aequorin triggered with Ca2+ (E). Curve C is a differential spectrum between aequorin and the protein residue (Shimomura et al., 1974b) protein concentration 0.5 mg/ml for A and B, 1.0 mg/ml for C. From Shimomura and Johnson, 1976.

Table 4.1.1 Main Properties of Natural Aequorin and its Ca2+-triggered Luminescence...

Activators and inhibitors. The total amount of light emitted in Ca2+-triggered luminescence is increased by certain alcohols for example, 10% by 2mM n-hexanol, 30% by 2mM n-heptanol, and 18% by saturated n-octanol (Shimomura et al., 1962 Neering and Fryer, 1986). The mechanism of the activation is unclear. No other types of activation is known. 

Fig. 4.1.15 Comparison of the luminescence and fluorescence emission spectra of natural aequorin (left panel) and recombinant e-aequorin (right panel) the luminescence spectra of Ca2+ -triggered reaction (dark solid lines), the fluorescence emission spectra of the spent solution containing 2 mM Ca2+ (dashed lines), and the luminescence spectra of the spent solution after addition of coelenterazine (light solid lines). Reproduced with permission, from Shimomura, 1995d. the Biochemical Society.

Preparation of semisynthetic aequorins. The best yield of semisynthetic aequorins can be obtained by using the apoaequorin prepared from aequorin immediately before use. Apoaequorin stored, even for 2-3 days, or recombinant apoaequorin isolated from a bacterial culture will give a significantly lower yield due to their somewhat unfolded molecular conformation. Fresh apoaequorin prepared by the Ca2+-triggered luminescence reaction appears to have the conformation best suited for regeneration. 

Fig. 4.2.1 Luminescence spectra of the Ca2+-triggered light emission of recombinant obelins (dotted lines), and the fluorescence emission spectra of their spent solution after luminescence (solid lines). Left obelin derived from O. geniculata right obelin derived from O. longissima. Reproduced from Markova etal., 2002, with permission from the American Chemical Society.

Concerning the Ca2+-triggered luminescence of obelin, Deng et al. (2001) reported an interesting observation (Fig. 4.2.2) the luminescence of the recombinant obelin from O. longissima is blue (7max 475 nm), whereas a mutant of this obelin, in which the amino acid residue-92 has been changed from tryptophan to phenylalanine, emits... 

Fig. 4.2.2 Left panel-. Uncorrected Ca2+-triggered bioluminescence spectrum of W92F obelin derived from O. longissima. Right panel Corrected bioluminescence spectrum of the same obelin (dotted line), and the fluorescence emission spectrum of the spent solution after luminescence (solid line). From Deng et al., 2001, with permission of the Federation of the European Biochemical Societies.

Fig. 4.8.4 Effect of pH on the light intensity of the Ca2+-triggered luminescence of mnemiopsin-1 (o), mnemiopsin-2 ( ), and berovin (A). From Ward and Seliger, 1974b, with permission from the American Chemical Society.

Links to phosphate signals Kinases and phosphatases with attached Ca2+ trigger proteins... 

Improved message system Growth connected to light. Ca2+ triggers... 

Extra-/Intra-cell communication Many novel Ca2+ trigger proteins (see Table 9.9)... 

The accompanying depolarization of the membrane at the synaptic ending permits a rapid inflow of calcium ions through a voltage-gated calcium channel.444 560 Within less than 0.1 ms the transient increase in intracellular [Ca2+] triggers the release of the contents of the vesicles. About four calcium ions are needed to release one clathrin-coated vesicle (Fig. 30-20A,B). The membrane fusion required for transmitter release involves cytoskeletal proteins of the synaptic endings as well as specialized proteins that are present in the membranes of the synaptic vesicles (Table 30-6). In fact, every step in the cycle depends upon specialized proteins.387... 

Factor VII is a single chain protein, which contains gla residues and shows N-terminal sequence homology with the other vitamin K-dependent blood proteins. Binding of Ca2+ triggers a conformational change which facilitates interaction with phospholipid.401... 

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