Kinases Subject

Smooth muscle contractions are subject to the actions of hormones and related agents. As shown in Figure 17.32, binding of the hormone epinephrine to smooth muscle receptors activates an intracellular adenylyl cyclase reaction that produces cyclic AMP (cAMP). The cAMP serves to activate a protein kinase that phosphorylates the myosin light chain kinase. The phosphorylated MLCK has a lower affinity for the Ca -calmodulin complex and thus is physiologically inactive. Reversal of this inactivation occurs via myosin light chain kinase phosphatase. 

AMP-activated protein kinases are heterotrimeric complexes comprised of catalytic a subunits and regulatory (3 and y subunits (Table 1). Each subunit is encoded by at least two genes, some of which can also be subject to alternate splicing, leading to a diverse array of possible heterotrimeric combinations. 

TRAM is subject to control through phosphorylation by protein kinase C-e. It is phosphorylated on serine 16 which is located close to the myristoylation site which is TRAM cannot signal without this phosphorylation or if the myristoylation site has been mutated. 

As mentioned above, many transcription factors are not always active. Rather the activity of transcription factors is often achieved by induced reversible modification. Most frequently is the addition of phosphate groups (phosphorylation) to Ser, Thr, or Tyr residues. For the AP-1 component c-Jun the phosphorylation at Ser63 and Ser73 enhances activity when cells are subjected to stress, e.g. radiation. Phosphorylation is, however, dispensable for c-Jun-dqDendent tissue homeostasis in the liver, indicating that certain activities do not require the regulatory enhancement. Jun-N-teiminal kinase and a kinase called RSK or p38 catalyze the phosphorylation of AP-1. 

Many proteins can be phosphorylated at multiple sites or are subject to regulation both by phosphorylation-dephosphorylation and by the binding of allosteric ligands. Phosphorylation-dephosphorylation at any one site can be catalyzed by multiple protein kinases or protein phosphatases. Many protein kinases and most protein phosphatases act on more than one protein and are themselves interconverted between active and inactive forms by the binding of second messengers or by covalent modification by phosphorylation-dephosphorylation. 

Recent evidence indicates that the 5-HT transporter is subject to post-translational regulatory changes in much the same way as neurotransmitter receptors (Blakeley et al. 1998). Protein kinase A and protein kinase C (PKC), at least, are known to be involved in this process. Phosphorylation of the transporter by PKC reduces the Fmax for 5-HT uptake and leads to sequestration of the transporter into the cell, suggesting that this enzyme has a key role in its intracellular trafficking. Since this phosphorylation is reduced when substrates that are themselves transported across the membrane bind to the transporter (e.g. 5-HT and fi -amphetamine), it seems that the transport of 5-HT is itself linked with the phosphorylation process. Possibly, this process serves as a homeostatic mechanism which ensures that the supply of functional transporters matches the demand for transmitter uptake. By contrast, ligands that are not transported (e.g. cocaine and the selective serotonin reuptake inhibitors (SSRIs)) prevent the inhibition of phosphorylation by transported ligands. Thus, such inhibitors would reduce 5-HT uptake both by their direct inhibition of the transporter and by disinhibition of its phosphorylation (Ramamoorthy and Blakely 1999). 

Ruiz, J.M. et ah. Proline metabolism and NAD kinase activity in green bean plants subjected to cold shock. Phytochemistry, 59, 473, 2002. 

VI. Valentine, W. N Tanaka, K. R., and Miwa, S A specific erythrocyte glycolytic enzyme defect (pyruvate kinase) in three subjects with congenital non-spherocytic hemolytic anemia. Trans. Assoc. Am. Physicians 74, 100-110 (1961). 

Multiple interactions are also being demonstrated between the traditional second-messenger pathways and the MAPK cascades. Free (3y G protein subunits, generated upon activation of receptors coupled to the G family, lead to activation of the ERK pathway. The mechanism by which this occurs, which may involve an interaction between the subunits and Ras or Raf, is a subject of intensive research (see Ch. 19). In addition, increases in cellular Ca2+ concentrations lead to stimulation of the ERK pathway, apparently via phosphorylation by CaMKs of proteins, for example She and Grb, that link growth factor receptor tyrosine kinases to Ras. Activation of the... 

RyRs do not exist as isolated SR ion channels, but as protein complexes subject to modulation by cellular metabolites, [Ca2+] , kinases and other factors (e.g. Marx et al 2000). Currently, very little is known with respect to the expression and localization of RyR isoforms, the regulatory factors underlying sensitivity of the complex to gating, and the effect of luminal (SR) Ca2+ on the probability of spontaneous or triggered release. A renewed focus on the role of RyRs in smooth muscle should help move the field from the initial discovery of these exciting phenomena to a clearer understanding of the function of this system in diverse smooth muscle tissues. 

Only three amino acids have a hydroxyl functional group in their side chain tyrosine, serine and threonine. Some kinases target only tyrosine residues (tyrosine kinases) whereas others may phosphorylate serine or threonine (Ser/Thr kinases). An enzyme protein (the substrate for the kinase) may have several tyrosine, serine or threonine residues within its primary sequence, but only some of these are subject to phosphorylation by a particular kinase (see Figure 3.6)... 

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