Phosphorylation hydrophobic association

Phosphorylation, the covalent attachment of a phosphate to an OH group, has been to date the most effective way to raise the temperature of the T,-divide. Thus, dephosphorylation, removal of phosphate, has been the most effective way to lower the temperature of the T,-divide and thereby the most effective way to drive hydrophobic association and its equivalent of contraction. This is similar to a primary event in muscle contraction (see Chapters 7 and 8). The shift in the T,-divide on binding of ATP can be as great as or greater than simple phosphorylation, depending on the interactions of ATP at the binding site. As discussed in Chapter 8, section 8.5, ATP binding drives hydrophobic dissociation, whereas loss of phosphate drives hydrophobic association both for the attachment to actin and for the power stroke to... 

Because hydrophobic association does not occur until the temperatiu e is above T, for the phosphorylated state, the phosphate cannot be folded into a low dielectric constant site, but instead would be fully exposed to the solvent. It is fundamental to the consilient mechanism of protein-based machines to understand how this can be As discussed in section 5.7, the high-energy state of the phosphate is due to competition for hydration between the phosphate and the hydrophobic groups, as reflected in the large change in Tt. 

It is important to note that tyrosine (Tyr, Y) and tryptophan (Tirp, W) are more hydrophobic than valine in the T,-based hydrophobicity scale. Therefore, Y and W would be candidates for the primary, the hydrophobic, family of the genetic code, except that these residues add additional physical properties. Both have large dipole moments and exhibit chemical reactivities that can dramatically change their oil-like character. Most notably, tyrosine can be a site for phosphorylation, which converts it to a supervinegar-like residue and dramatically disrupts hydrophobic folding, assembly, and associated functions. On our scale, based directly on the hydrophobic association event, tryptophan is the most hydrophobic residue, whereas other scales that utilize less direct means of assessing functional hydrophobicity place trytophan at much less hydrophobic positions. Thus, these residues, tyrosine and tryptophan, would not... 

As calculated and discussed in section S.3.4.4.4, the AGha(P04") due to phosphorylation of poly[30(GVGIP)(GRGDSP)] at the Ser (S) residue raises the value of T, to an extent that the calculated increase in free energy for hydrophobic association becomes about... 

GVGIP)n. Phosphorylation removes more than two-thirds of the proton-proton contacts that report the hydrophobic associations within the p-barrel. 

As is apparent in Chapter 8, phosphates are the most polar molecular species available in biology for controlling hydrophobic association/ dissociation, that is, for controlling processes that occur by inverse temperature transition. It follows, therefore, that kinases for phosphorylation and phosphatases for dephosphorylation would be fundamental to key cellular trans-ductional and transformational processes. Often in cancerous and other diseased states, the activities of these enzymes are abnormal. Importantly for our interests, their sites of interaction can be very selective. Changes in protein kinase C activities have been reported to be abnormal in colon, breast, and skin cancers. Protein tyrosine kinase, which selectively phos-phorylates the internal tyrosine (Tyr,Y) residue in the sequence GIYWHHY, is overexpressed in colon and breast cancer. " Furthermore, cyclic AMP-dependent protein kinase activities have been associated with the onset of... 

Relevance of Gibbs Free Energy for Hydrophobic Association to Energizing by Phosphorylation... 

FIGURE 8.8 Mechanism of activation of protein kinase B (PKB). PI3-kinase is recruited to the membrane via direct association with the receptor PTK or via association with the docking protein Gab-1. It catalyzes the generation of phosphatidyl-3,4,5-inositolphosphate, which serves as a membrane-recruitment signal for PKB. Associated with the membrane, it is first phosphorylated in its catalytic domain by PDK1 and then by PDK2 in the hydrophobic motif. The activated PKB then detaches from the membrane. 

N-Myristoylation is achieved by the covalent attachment of the 14-carbon saturated myristic acid (C14 0) to the N-terminal glycine residue of various proteins with formation of an irreversible amide bond (Table l). 10 This process is cotranslational and is catalyzed by a monomeric enzyme called jV-myri s toy 11ransferase. 24 Several proteins of diverse families, including tyrosine kinases of the Src family, the alanine-rich C kinase substrate (MARKS), the HIV Nef phosphoprotein, and the a-subunit of heterotrimeric G protein, carry a myr-istoylated N-terminal glycine residue which in some cases is in close proximity to a site that can be S-acylated with a fatty acid. Functional studies of these proteins have shown an important structural role for the myristoyl chain not only in terms of enhanced membrane affinity of the proteins, but also of stabilization of their three-dimensional structure in the cytosolic form. Once exposed, the myristoyl chain promotes membrane association of the protein. 5 The myristoyl moiety however, is not sufficiently hydrophobic to anchor the protein to the membrane permanently, 25,26 and in vivo this interaction is further modulated by a variety of switches that operate through covalent or noncovalent modifications of the protein. 4,5,27 In MARKS, for example, multiple phosphorylation of a positively charged domain moves the protein back to the cytosolic compartment due to the mutated electrostatic properties of the protein, a so-called myristoyl-electrostatic switch. 28 ... 

VR1 is a polytopic protein containing six transmembrane segments with an additional short hydrophobic stretch between transmembrane regions 5 and 6, which is believed to be associated with the channel pore. There are three possible protein kinase A phosphorylation sites on the VR1 that might play a role in receptor desensitisation. 

A hydrophobic 10 kDa protein is also associated with PS II preparations [14], but its function is obscure. The protein is phosphorylated by a membrane-bound protein kinase [15] and the identity of this phosphoprotein has been the subject of much speculation [16]. It is clearly not the 9 kDa polypeptide of Cyt 6-559 or the 8 kDa proteolipid subunit of ATP synthase as shown by N-terminal amino acid sequence [14]. 

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