Phospholipase, domain structure

Phospholipases. Figure 3 Representation ofthe domain structure of phospholipase D1 (adapted from [4]). 

Figure 15.11. Modular Structure of Phospholipase C. The domain structures of three isoforms of phospholipase C reveal similarities and differences among the isoforms. Only the P isoform, with the G-protein-binding domain, can be stimulated directly by G proteins. For phospholipase Cy, the insertion of two SH2 (Src homology 2) domains and one SH3 (Src homology 3) domain splits the catalytic domain and a PH domain into two parts.

These phospholipases are primarily involved in signal transduction. The structure and mechanism of action of phosphatidylinositol-specific PLCs from both manunalian and bacterial sources are reviewed in Refs. [31,32]. The structure of manunalian phos-phatidylinositol-PLC5 is shown in Fig. 10 and highlights the domain structure. This protein was crystallized in the absence of its PH domain that is at the N-terminus. A tether and fix model of membrane association is suggested (L.-O. Essen, 1996) whereby initial... 

Ferguson, K.M., et al. Structure of the high affinity complex of inositol triphosphate with a phospholipase C pleckstrin homology domain. Celt 83 1037-1046, 1995. 

Perisic, O., Fong, S., Lynch, D.E., Bycroft, M., and WUhams, R.L., 1998, Crystal structure of a calcium-phosphohpid binding domain from cytosohc phospholipase A2. J. Biol. Chem. 273 1596-1604. 

Phosphohpases of type Cy are activated by receptor tyrosine kinases (see Chapter 8), and thus phosphohpase Cy is involved in growth factor controlled signal transduction pathways. The receptor tyrosine kinases (see Chapter 8) phosphorylate the enzyme at specific tyrosine residues and initiate activation of the enzyme. Characteristic for the structure of phospholipase Cy is the occurrence of SH2 and SH3 domains (see Chapter 8). These represent protein modules that serve to attach further partner proteins. 

Fig. 8.11. Recognition of phosphotyrosine-containing substrate peptides by SH2 domains of Src kinase and phospholipase Cyl. Binding of phosphotyrosine-containing peptides to SH2 is shown schematically, based on crystal structures of the complexes. The SH2 domain of Src kinase has a basic binding pocket for the phosphotyrosine residue and a hydrophobic pocket for the isoleucine residues at position -1-3 of the peptide substrate. The SH2 domain of PL-Cyl has a hydrophobic binding surface to which the C-terminal part of the peptide P-Tyr-Ile-Ile-Pro-Leu-Pro-Asp binds. According to Cohen, (1995).

The pleckstrin homology (PH) domain is a structural motif of ca. 100 amino acids foimd in many signal molecules such as Ser/Thr-specific protein kinases, tyrosine kinases, isoforms of phospholipase C (PL-CP, y and 6), in G nucleotide exchange factors, adaptor proteins, and in proteins of the cytoskeleton (see also Fig. 8.10). Originally, the PH domain was foimd in the 47 kDa pleckstrin protein, which is the main substrate of protein kinase C in platelets. 

Domain III shares no sequence homology with other known proteins, its structure resembling the C2 domains found in phospholipase C, protein kinase C and synap-totagmin (Rizo and Sudhof, 1998). In addition to linking the Ca2+-binding domain of the molecule to the catalytic domain (domain II), domain III appears to be involved in phospholipids and Ca2+ binding (Tompa et al., 2001). 

During last decades the domains C-2 symmetry (the dyad rotation symmetry) of low-B palindrome was established in many enzymes (chymotrypsin, trypsin, aspartyl proteinases, HIV-1 protease, carboxypeptidase A, phospholipase A-2 ribonuclease, etc.) (Lumry, 2002 and references therein). It is proposed that the pair domain closure causes constrain of pretransition state complex that activates cleavage or formation of chemical bonds. Thus control of strong bonds by the cooperation of many matrix or knots bonds takes place. As an example, in the active site of carboxypeptidase A the zinc ion is attached to one of the catalytic domains by histidine 69 and glutamine 72 and connected by hystidine 196 to the second domain. Similar structures were found in the chymotrypsin and pepsin active sites where protons are driven under compression of the domains closure. 

The three-dimensional structure of NTE has not been experimentally determined, but a homology model of the patatin domain indicates that the active site serine (Ser ) is located on a nucleophilic elbow characteristic of serine hydrolases (Wijeyesakere et al., 2007). Moreover, the model indicates that the catalytic site of NTE consists of a novel Ser-Asp catalytic dyad, as in patatin and mammalian cytosolic phospholipase A2 (CPLA2), rather than the classical catalytic triad (Ser-Asp/Glu-His), as found in many serine hydrolases including AChE. Recently, mutations have been identified in NTE that are associated with motor neuron disease (Rainier et al., 2008). The mutations occur within the catalytic domain of NTE, but it is not yet known if they affect the catalytic function of the enzyme or alter some other property of the protein in order to produce disease. 

With this structure in mind, we can now understand how PKC is activated on PIP2 hydrolysis (Figure 15.16B). Before activation, PKC is free in solution. On PIP2 hydrolysis in the membrane by phospholipase C, the CIB domain of PKC... 

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