ADP Uptake

Electrogenic antiport, ADP uptake favored by the membrane potential can also mediate electroneutral exchange of ADPout for... 

Note These are examples of important transporters involved in substrate and ADP uptake into the matrix compartment as indicated, and most are reversible. These transporters are proteins and several have been isolated and sequenced. Other specific carriers occur in mitochondria from other tissues. The inner membrane does not allow rapid exchange of NAD or CoA but there are mechanisms for the slow uniport of cofactors synthesized extramitochondrially. 

Table 7.2. Effects of various metabolites on [a32P]ADP uptake into E. coli cells expressing several adenine nucleotide carriers. For uptake experiments, effectors were always present in a fivefold higher concentration than the given uptake substrates. [a32P]ADP uptake by AAC2(A.t.), AACl(N.sp.), HMP31(T.g.), and ANTl( ./z.) was measured at a substrate concentration of 10, 100, 50, and 200 pM, respectively (Voncken et al. 2002 Tjaden et al. 2004 Leroch et al. 2005 Leroch 2006) ...

FIGURE 8 ATP, ADR and Pj transport in mitochondria. ATP is formed inside mitochondria. Most of the ATP is exported to the cytoplasm where it is cleaved to ADP and Pj. The mitochondrial inner membrane contains specific proteins that mediate not only ATP release coupled to ADP uptake, but also Pj uptake linked to hydroxide ion (OH ) release. 

Adenosine is formed from ATP via a phosphatase cascade that sequentially involves the diphosphate, ADP, and the monophosphate, AMP. The actions of adenosine are terminated by uptake and rephosphorylation via adenosine kinase to AMP or by cataboHsm via adenosine deaminase to inosine and hypoxanthine. 

Fructose is present outside a cell at 1 /iM concentration. An active transport system in the plasma membrane transports fructose into this cell, using the free energy of ATP hydrolysis to drive fructose uptake. Assume that one fructose is transported per ATP hydrolyzed, that ATP is hydrolyzed on the intracellular surface of the membrane, and that the concentrations of ATP, ADP, and Pi are 3 mM, 1 mM, and 0.5 mM, respectively. T = 298 K. What is the highest intracellular concentration of fructose that this transport system can generate Hint Kefer to Chapter 3 to recall the effects of concentration on free energy of ATP hydrolysis.)... 

Chemical modification studies with fluorescein-5 -isothiocyanate support the proximity of Lys515 to the ATP binding site [98,113-117,212,339]. Fluorescein-5 -isothiocyanate stoichiometrically reacts with the Ca -ATPase in intact or solubilized sarcoplasmic reticulum at a mildly alkaline pH, causing inhibition of ATPase activity, ATP-dependent Ca transport, and the phosphorylation of the Ca " -ATPase by ATP the Ca uptake energized by acetylphosphate, carbamylphos-phate or j -nitrophenyl phosphate is only partially inhibited [113,114,212,339]. The reaction of -ATPase with FITC is competitively inhibited by ATP, AMPPNP, TNP-ATP, and less effectively by ADP or ITP the concentrations of the various nucleotides required for protection are consistent with their affinities for the ATP binding site of the Ca -ATPase [114,212,340]. 

A well-known example of active transport is the sodium-potassium pump that maintains the imbalance of Na and ions across cytoplasmic membranes. Flere, the movement of ions is coupled to the hydrolysis of ATP to ADP and phosphate by the ATPase enzyme, liberating three Na+ out of the cell and pumping in two K [21-23]. Bacteria, mitochondria, and chloroplasts have a similar ion-driven uptake mechanism, but it works in reverse. Instead of ATP hydrolysis driving ion transport, H gradients across the membranes generate the synthesis of ATP from ADP and phosphate [24-27]. 

Figure 3. Examples of major types of uptake mechanisms realised in prokaryotic outer membranes (a to c) and cytoplasmic membranes (a, and d to 1). The solutes to be transported are shown by filled circles x symbolises another solute which is transported in the same or in the opposite direction. In systems h-k, uptake is driven by the cleavage of ATP to ADP and phosphate. One type of uptake system, 1, depends on the energy-rich molecule phosphoenolpyruvate shown as PEP .

It was also observed in earlier studies that mitochondria not only accumulate Ca2+ as an alternative to phosphorylation of ADP (Ca2+ uptake uncouples phosphorylation from electron transport), but could also accumulate much larger amounts of Ca2+ if phosphate was also taken up, resulting in precipitation of Ca2+ within the matrix as insoluble hydroxyapatite, visible as electron-dense granules by EM. An unusual feature of these hydroxyapatite deposits is that they fail to become crystalline and remain amorphous even over protracted periods of time. Their presence in mitochondria in a number of disease conditions underlines the role for mitochondria as a sort of safety device, which can enable the cell to survive, if only for a limited period of time, situations of cytoplasmic Ca2+ overload. 

A different and simpler approach to the measurement of P/O ratios came from the introduction of an oxygen electrode suitable for biochemical studies. Chance and Williams (1955) established conditions under which mitochondrial respiration, in the presence of excess substrate, was totally dependent on the amount of ADP available, i.e., the mitochondria were exhibiting respiratory control. From the change in potential when a known amount of ADP was admitted into the electrode vessel, the oxygen uptake and thus the P/O ratio could be determined, completely confirming the earlier results. 

Barth H, Blocker D, Aktories K. The uptake machinery of clostridial actin ADP-ribosylating toxins—a cell delivery system for fusion proteins and polypeptide drugs. Naunyn Schmiedebergs Arch Pharmacol 2002 366(6) 501-512. 

The binary nature of iota toxin from C. perfringens type E was first explored in 1986 by Stiles and Wilkins. The overall mode of action for iota toxin is widely comparable to C2 toxin. The binding/translocation component iota b (Ib) facilitates cellular uptake of the enzyme component iota a (la) in a like manner as previously described for C2 toxin. la, just as C2I, specifically mono-ADP-ribosylates G-actin at Argl77. ... 

Glucose is an essential fuel for the brain and, if the blood concentration falls, uptake by the brain decreases and less fuel is available for ATP generation in the neurones. This results in a decrease in the ATP/ADP concentration ratio. Consequently, less energy is released on ATP hydrolysis, so that less is available for synthesis, transport of neurotransmitter within the nerve and release into the synapse. Hypoglycaemia could, therefore, reduce the effectiveness of neurotransmitters which would reduce stimulation of the motor control pathway. The result would be inhibition of muscle contraction (Figure 13.27). 

Four different stages can be distinguished in the enzyme s catalytic cycle (2). First, binding of ATP to the N domain leads to the uptake of two Ca " into the transmembrane part (a). Phosphorylation of an aspartate residue in the P domain (b) and dissociation of ADP then causes a conformation change that releases the Ca " ions into the SR (c). Finally, dephosphorylation of the aspartate residue restores the initial conditions (d). 

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