MSos protein

The structure of the mSos protein (m=mammalian) is shown in Fig. 9.8. Sequence comparison with known Ras-specific GEFs has identified a common domain of ca.200 amino acids, to which nucleotide exchange activity has been assigned. Within this domain, three highly conserved sequence elements can be differentiated, separated by more variable sections. Other structural elements include a PH domain and a Pro-rich binding domain. The Pro-rich sequence fimctions as an attachment site for the SH3 group of Grb2 protein. 

The complex of Grb2 and mSos proteins forms a link between Ras protein and activated receptor tyrosine kinases. The Grb2 protein has two SH2 domains (see 8.5) and a SH3 domain. 

The insulin receptor substrate IRS couples the insulin receptor to sequential effector molecules (review Ogawa et al., 1998). On binding of insulin to the insulin receptor, the tyrosine kinase activity of the receptor is stimulated. The IRS protein is phosphory-lated at several Tyr residues, which then serve as attachment points for sequential effector molecules as e.g. the Grb2-mSos complex, the P13-kinase and the protein tyrosine phosphatase SHP-2. The IRS protein also has a phosphotyrosine binding domain and a PH domain. Both modules are required for signal transduction in vivo. It is assumed that the PTB domain binds to autophosphorylation sites of the insulin receptor and that the PH domain is involved in membrane association of IRS. 

Fig. 9.8. Domain structure of mSos. The mSos-1 protein of mammals possesses a pleckstrin homology domain (PH), a Pro-rich domain for interaction with Grb2 and a catalytic domain with three sequence motifs (1,2,3) characteristic for Ras GEFs.

This well characterized pathway of Ras signal transmission was the first to be discovered (see above) and involves adaptor proteins (Grb2, She) and GEFs (e.g. mSos). 

Fig. 9.9 Model of the function of Grb2-mSos and the adaptor protein She in the Ras pathway. The figure shows a highly simplified version of the two known pathways of involvement of the Grb2-mSos complex in signal transduction via the Ras protein. Phosphotyrosine residues of an activated, auto-phosphorylated receptor R may serve as attachment points forthe PTB domains of the She adaptor protein (a) or forthe SH2 domain of the Grb2-mSos...

Inhibitor studies (Leaet al., 1979) have shown that the synthesis of protein from glutamine by isolated pea cotyledons is unaffected by MSO but inhibited by azaserine, consistent with the action of glutamate synthase but not very conclusive because of the side effects of azaserine. Little if any labeling data on ammonia assimilation has so far been produced using developing seeds. 

Table 3. Peak list comparison. List of masses showing a difference of "n x 14.0157 Da" with less than 15 PPM error between MALDl-MS spectra from untreated and esterified samples. Init. mass list of masses in the native sample spectrum that matched with peak masses of the esterified sample. Mass ester masses in the esterified sample spectrum ester potential number of ester groups on the peptide chain (n from 1 to 6) D E potential number ofD and E amino acids in the peptide sequence (n-1 due to the C-termirutl esterification) Protein source the protein to which the peptide corresponds (Protein named 095678 and K2C1 HUMAN are two cytoskeletal keratin type II from hair) Bold letters in sequence column represent potential position of the esterification site on the sequence from D and E amino acids and letters in italics represent potential esterification at the C-terminal amino acid. MSO methionine sulfoxide...

Several inhibitors of the GS (Methionine sulphoximine, MSO Phos-phinothricin, PPT) have been used in studies of the role of this key and the mechanism of its action. Both substances can inhibit GS "in vivo", cause a dramatic increase of ammonia and inhibit the rate of photosynthetic CO2 uptake. Initially it was thought that the build-up of ammonia was the cause of photosynthetic inhibition however, Sauer et al. (1987) postulated that ami noacid metabolism, disturbed by PPT, results in a) Inhibition of synthesis of proteins.e.g. the Qb protein causing eventually the collapse of electron transport ... 

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