Modified amino-terminal

Recombinant DNA technology can also be used to create modified proteins that are readily purified by affinity chromatography. The gene of interest is Unked to an oligonucleotide sequence that encodes a carboxyl or amino terminal extension to the encoded protein. The... 

Figure 46-8. Fusion of a vesicle with the plasma membrane preserves the orientation of any integral proteins embedded in the vesicle bilayer. Initially, the amino terminal of the protein faces the lumen, or inner cavity, of such a vesicle. After fusion, the amino terminal is on the exterior surface of the plasma membrane. That the orientation of the protein has not been reversed can be perceived by noting that the other end of the molecule, the carboxyl terminal, is always immersed in the cytoplasm. The lumen of a vesicle and the outside of the cell are topologically equivalent. (Re drawn and modified, with permission, from Lodish HF, Rothman JE The assembly of cell membranes. Sci Am [Jan] 1979 240 43.)...

Proudfoot AE, Buser R, Borlat F et al (1999) Amino-terminally modified RANTES analogues demonstrate differential effects on RANTES receptors. J Biol Chem 274 32478-32485 Qin AP, Zhang HE, Qin ZH (2008) Mechanisms of lysosomal proteases participating in cerebral ischemia-induced neuronal death. Neurosci Bull 24 117-123 Richter R, Bistrian R, Escher S et al (2005) Quantum proteolytic activation of chemokine CCL15 by neutrophil granulocytes modulates mononuclear cell adhesiveness. J Immunol 175 1599-1608... 

Nihonyanagi, S., Ye, S. and Uosaki, K. (2001) Sum frequency generation study on the molecular structures at the interfaces between quartz modified with amino-terminated self-assemhled monolayer and electrolyte solutions of various pH and ionic strength. Electrochim. Acta, 46, 3057—3061. 

Chvatchko Y, Proudfoot AE, Buser R, et al. Inhibition of airway inflammation by amino-terminally modified RANTES/CC chemokine ligand 5 analogues is not mediated through CCR3. J Immunol 2003 171(10) 5498-5506. 

Yeung, C.W.T., Moule, M.L., and Yip, C.C. (1980) Photoaffinity labeling of insulin receptor with an insulin analogue selectively modified at the amino terminal of the B chain. Biochemistry 19, 2196-2203. 

Chromatin-modifying complexes are classified into two major groups (1) enzymes that conttol covalent modifications of the amino-terminal tails of histones (acetylation, methylation, phosphorylation, ubiquitinylation) (see Sections 1.3 and... 

Amino-terminated organosilanes were used to immobilize rifamycin B via its active carboxylic acid functionality leading to the formation of stable amide bonds between antibiotics and modified silica [7]. The organosilanes used were (3-aminopropyl)triethoxysilane or (3-aminopropyl)dimethylethoxysilane, and the procedure for preparation was as described in Section 2.2.1.1. Surface coverage data were not provided by the authors [7],... 

Identification of the N-terminal Amino Acid in Polypeptides (TLC of Modified Amino Acids)... 

Amino-Terminal and Carboxyl-Terminal Modifications The first residue inserted in all polypeptides is Al-formylmethio-nine (in bacteria) or methionine (in eukaryotes). However, the formyl group, the amino-terminal Met residue, and often additional amino-terminal (and, in some cases, carboxyl-terminal) residues may be removed enzymatically in formation of the final functional protein. In as many as 50% of eukaryotic proteins, the amino group of the amino-terminal residue is Al-acetylated after translation. Carboxyl-terminal residues are also sometimes modified. 

Source Modified from Bachmair, A., Finley, D., Varshavsky, A. (1986) In vivo half-life of a protein is a function of its amino-terminal residue. Science 234, 179-186. 

Half-lives were measured in yeast for the j8-galactosidase protein modified so that in each experiment it had a different amino-terminal residue. (See Chapter 9 for a discussion of techniques used to engineer proteins with altered amino acid sequences.) Half-lives may vary for different proteins and in different organisms, but this general pattern appears to hold for all organisms. 

Histones within transcriptionally active chromatin and heterochromatin also differ in their patterns of covalent modification. The core histones of nucleosome particles (H2A, H2B, H3, H4 see Fig. 24-27) are modified by irreversible methylation of Lys residues, phosphorylation of Ser or Thr residues, acetylation (see below), or attachment of ubiquitin (see Fig. 27-41). Each of the core histones has two distinct structural domains. A central domain is involved in histone-histone interaction and the wrapping of DNA around the nucleosome. A second, lysine-rich amino-terminal domain is generally positioned near the exterior of the assembled nucleosome particle the covalent modifications occur at specific residues concentrated in this amino-terminal domain. The patterns of modification have led some researchers to propose the existence of a histone code, in which modification patterns are recognized by enzymes that alter the structure of chromatin. Modifications associated with transcriptional activation would be recognized by enzymes that make the chromatin more accessible to the transcription machinery. 

The pathway of protein synthesis translates the three-letter alphabet of nucleotide sequences on mRNA into the twenty-letter alphabet of amino acids that constitute proteins. The mRNA is translated from its 5 -end to its 3 -end, producing a protein synthesized from its amino-terminal end to its carboxyl-terminal end. Prokaryotic mRNAs often have several coding regions, that is, they are polycistronic (see p. 420). Each coding region has its own initiation codon and produces a separate species of polypeptide. In contrast, each eukaryotic mRNA codes for only one polypeptide chain, that is, it is monocistronic. The process of translation is divided into three separate steps initiation, elongation, and termination. The polypeptide chains produced may be modified by posttranslational modification. Eukaryotic protein synthesis resembles that of prokaryotes in most details. [Note Individual differences are mentioned in the text.]... 

Other modified amino acids have been used as y-turn inducers. For example, Toniolo et al. 21 utilized a reduced peptide isostere (-CH2NH-) moiety and incorporated it into the neurotransmitter tripeptide -Pro-Leu-Gly-. The X-ray diffraction structure of the N-terminal Boc-or Z-protected tripeptide Boc(or Z)-Prot i[CH2NH]Leu-Gly-NH2 2 provides unequivocal proof of the existence of a y-turn (Scheme 3). 

Copolymers. Copolymers from mixtures of different bisphenols or from mixtures of dichlorosulfone and dichlorobenzophenone have been reported in the patent literature. Bifunctional hydroxyl-terminated polyethersulfone oligomers are prepared readily by the polyetherification reaction simply by providing a suitable excess of the bisphenol. Block copolymers are obtained by reaction of the oligomers with other polymers having end groups capable of reacting with the phenol. Multiblock copolymers of BPA-polysulfone with polysiloxane have been made in this way by reaction with dimethyl amino-terminated polydimethylsiloxane the products are effective impact modifiers for the polyethersulfone (79). Block copolymers with nylon-6 are obtained when chlorine-terminated oligomers, which are prepared by polyetherification with excess dihalosulfone, are used as initiators for polymerization of caprolactam (80). 

The COOH-terminal amino acid of a peptide or protein may be analyzed by either chemical or enzymatic methods. The chemical methods are similar to the procedures for NH2-terminal analysis. COOH-terminal amino acids are identified by hydrazinolysis or are reduced to amino alcohols by lithium borohydride. The modified amino acids are released by acid hydrolysis and identified by chromatography. Both of these chemical methods are difficult, and clear-cut results are not readily obtained. The method of choice is peptide hydrolysis catalyzed by carboxypeptidases A and B. These two enzymes catalyze the hydrolysis of amide bonds at the COOH-terminal end of a peptide (Equation E2.3), since carboxypeptidase action requires the presence of a free a-carboxyl group in the substrate. 

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