Replacement modifications proteins

Fat Content of Milk. Raw milk as well as many dairy products are routinely analyzed for their fat content. The Babcock test, or one of its modifications, has been a standard direct measure for many years and is being replaced by indirect means, particularly for production operations. The Babcock test employs a bottle with an extended and caHbrated neck, milk plus sulfuric acid [7664-93-9] to digest the protein, and a centrifuge to concentrate the fat into the caHbrated neck. The percentage of fat in the milk is read direcflv from the neck of the bottle with a divider or caHper, rea ding to... 

Process Va.ria.tlons. The conventional techniques for tea manufacture have been replaced in part by newer processing methods adopted for a greater degree of automation and control. These newer methods include withering modification (78), different types of maceration equipment (79), closed systems for fermentation (80), and fluid-bed dryers (81). A thermal process has been described which utilizes decreased time periods for enzymatic reactions but depends on heat treatment at 50—65°C to develop black tea character (82). It is claimed that tannin—protein complex formation is decreased and, therefore, greater tannin extractabiUty is achieved. Tea value is beheved to be increased through use of this process. 

The determination of the primary structure of a protein is a very demanding analytical task, but, thanks to automated procedures, many of these structures are now known. Any modification of the primary structure of a protein—the replacement of one amino acid residue by another—may lead to a congenital disease. Even one wrong amino acid in the chain can disrupt the normal function of the molecule (Fig. 19.18). 

The second method also relies on site-specific chemical modification ofphosphoproteins (Oda et al., 2001). It involves the chemical replacement of phosphates on serine and threonine residues with a biotin affinity tag (Fig. 2.7B). The replacement reaction takes advantage of the fact that the phosphate moiety on phosphoserine and phosphothreonine undergoes -elimination under alkaline conditions to form a group that reacts with nucleophiles such as ethanedithiol. The resulting free sulfydryls can then be coupled to biotin to create the affinity tag (Oda et al., 2001). The biotin tag is used to purify the proteins subsequent to proteolytic digestion. The biotinylated peptides are isolated by an additional affinity purification step and are then analyzed by mass spectrometry (Oda et al., 2001). This method was also tested with phosphorylated (Teasein and shown to efficiently enrich phosphopeptides. In addition, the method was used on a crude protein lysate from yeast and phosphorylated ovalbumin was detected. Thus, as with the method of Zhou et al. (2001), additional fractionation steps will be required to detect low abundance phosphoproteins. 

The principal difference in the overall protein composition of PNS and CNS myelin is that P0 replaces PLP as the major protein, although myelin-forming Schwann cells do express very low levels of PLP. It is interesting to note that PLP and P0 proteins, which are so different in sequence, post-translational modifications and membrane topology, may have similar roles in the formation of structures as closely related as myelin of the CNS and PNS respectively. Expression of P0 in transfected cells results in cell-cell interactions that are due to homophilic binding... 

By means of this method, a variety of Ras proteins with different lipidation patterns could be synthesized in multimilligram amounts. For instance, proteins were generated with the natural lipid combination, i.e. a farnesyl thioether and a palmitoyl thioester. Furthermore, analogous proteins were synthesized embodying only one lipid residue in which either the farnesyl- or the palmitoyl group was replaced by a stable hexadecyl thioether. In addition, proteins were built up containing a serine instead of a cysteine residue at the critical sites which normally are lipidated. In a further series of experiments, lipidated Ras proteins which carry a fluorescent Mant group incorporated into the farnesyl-type modification were synthesized.1251... 

Replacement of the famesyl group by lipid analogues could be performed for full length Ras proteins in vitro by means of the enzyme famesyltrans-ferase. When such partially modified Ras constructs were applied in Xenopus oocytes the cellular machinery completed modification (endoprotease activity, carboxymethylation and palmitoylation). In these cases the H-Ras famesyl group could be stripped off most of its isoprenoid features that distinguish it from a fatty add without any apparent effect on its ability to induce oocyte maturation and activation of mitogen-activated protdn kinase In contrast, replacement by the less hydrophobic isoprenoid geranyl causes severely delayed oocyte activation. 

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