Amino acid sequence, basic protein from

Since the native conformation of a protein is ultimately determined by its primary sequence and its intracellular environment, it is theoretically possible to predict this native 3-dimensionaI structure from the amino acid sequence. Basically, this amounts to determining the minimum energy conformation of a protein, an approach that is still in its early stages of development, and which is not sufficiently advanced to allow reliable predictions of native protein structures. 

To note, intermediate filaments are cytoskeletal structures that are formed by different members of a family of related, highly conserved proteins (Godsel et al. 2008). Most types of intermediate filaments are located in the cytoplasm. The nuclearly localized intermediate filaments are called lamins. Categorization of intermediate filaments into six groups has been done on the basis of similarities in amino acid sequence and protein structure. Types I and II intermediate filaments are acidic and basic keratins, namely epithelial keratins and trichocytic keratins (e.g., in hair and horns). Type III intermediate filaments are, e.g., vimentin (widely expressed in fibroblasts), Type IV intermediate filaments are, e.g., neurofilaments, type V intermediate filaments are nuclear lamins, type VI intermediate filaments are nestin. A well-known disease resulting from... 

Biosynthesis. CRE is derived from a precursor of 196 amino acids (84,85). This gene contains one copy of CRE, which is flanked by double basic amino acids. The amino acid sequence of the CRE precursor suggests that it may arise from proteins related to POMC and neurophysins (31). The CRE precursor contains a cAMP responsive element which aHows stimulation of mRNA synthesis when intraceHular levels of cAMP are increased (86). 

To understand the biological function of proteins we would therefore like to be able to deduce or predict the three-dimensional structure from the amino acid sequence. This we cannot do. In spite of considerable efforts over the past 25 years, this folding problem is still unsolved and remains one of the most basic intellectual challenges in molecular biology. 

Separation of the purified PL1, PL2 and PL3 isoenzymes by SDS-PAGE yielded single protein bands that corresponded to a molecular mass of 42 kDa (6), 4 kDa higher than calculated from the deduced amino acid sequences. Isoelectric focusing revealed a pi of >10 for each isoenzyme similar to that of the basic Ech PLs (15). 

Lipid transfer peptides and proteins occur in eukaryotic and prokaryotic cells. In vitro they possess the ability to transfer phospholipids between lipid membranes. Plant lipid transfer peptides are unspecific in their substrate selectivity. They bind phosphatidylcholine, phosphatidylethanolamine, phosphatidylinositol, and glycolipids. Some of these peptides have shown antifungal activity in vitro The sequences of lipid transfer proteins and peptides contain 91-95 amino acids, are basic, and have eight cysteine residues forming four disulfide bonds. They do not contain tryptophan residues. About 40% of the sequence adopts a helical structure with helices linked via disulfide bonds. The tertiary structure comprises four a-helices. The three-dimensional structure of a lipid transfer peptide from H. vulgare in complex with palmitate has been solved by NMR. In this structure the fatty acid is caged in a hydrophobic cavity formed by the helices. 

Small ACTH fragments related to ACTH-(4-10) have also been investigated for the presence of ordered structure. CD of ACTH--(5-10) in TFE showed a random structure (50) as was found with H-NMR for fragment 4-10 (51). The addition of anionic or cationic surfactants to an aqueous solution of ACTH-(4-11) dit not promote any a-helix or 3-form in this peptide (CD experiments S2). When ACTH-(1-14) and 1-10 were measured by CD and NMR respectively, indications for a helical or ordered structure were found (90, ). Thus it seems that the addition of the non-helix "prone" fragment 1-3 or 1-4 can promote the formation of a helical structure in the adjacent sequence. Arguments in favour of this come from the theoretical work of Argos and Palau (53) on amino acid distribution in protein secondary structures. They found that Ser and Thr frequently occur at the N-terminal helical position (cf. Ser in ACTH) to provide stability the position adjacent to the helical C-terminus is often occupied by Gly or Pro (adjacent toTrp in ACTH we have Gly ) acidic amino acid residues are frequently found at the helix N-terminus (cf. Glu in ACTH) and/or basic residues at the C-terminus (cf. Arg ). 

Many secreted proteins, as well as smaller peptide hormones, are acted upon in the endoplasmic reticulum by tryptases and other serine proteases. They often cut between pairs of basic residues such as KK, KR, or RR.214-216 A substilisin-like protease cleaves adjacent to methionine.217 Other classes of proteases (e.g., zinc-dependent carboxypeptidases) also participate in this processing. Serine carboxypeptidases are involved in processing human prohormones.218 Among the serine carboxypeptidases of known structure is one from wheat219 and carboxypeptidase Y, a vacuolar enzyme from yeast.220 Like the pancreatic metallocarboxypeptidases discussed in Section 4, these enzymes remove one amino acid at a time, a property that has made carboxypeptidases valuable reagents for determination of amino acid sequences. Carboxypeptidases may also be used for modification of proteins by removal of one or a few amino acids from the ends. 

Not only this heptapeptide but also the entire amino acid sequence of a-melanotropin is found within the sequence of corticotropin (Fig. 30-2), which has an additional 29 amino acids at the C-terminal end.25 The same heptapeptide was also found in the lipotropins. The explanation is, in part, that several of these hormones arise from a single 31-kDa precursor protein called prepro-opiomelanocortin.25 26 It contains an N-terminal signal sequence that is removed shortly after synthesis, as well as pairs of adjacent basic residues (Arg-Arg, Arg-Lys, Lys-Arg and Lys-Lys) at a number of places (Fig. 30-2). After removal of the... 

Another useful structure tool is RasMol (or RasMac). This will allow you to view the detailed structure of a protein and rotate it on coordinates so you can see it from all perspectives. A hyperlink to RasMol is present under the View Structure function just above Chime. You may need to study RasMol instructions provided under Help, or you may use a Ra.s Mol tutorial listed in Table El.2. Another useful protein viewer is tin-Swiss-Protein Pdv Viewer (Table El.2). BLAST is an advanced sequence similarity tool available at NCBI. To access this, go to the NCBI home page (www.ncbi.nlm.nih.gov) and click on BLAST. Then click on Basic BLAST search to obtain a dialogue box into which you may type the amino acid sequence of human a-lactalbumin. This process may be stream lined by downloading the amino acid sequence in FASTA format into a file and transferring the fde into the BLAST dialogue box. BLAST will provide a list of proteins with sequences similar to the one entered. 

Fig. 6. Amino acid sequence similarity between the three G-box-binding proteins isolated from Arabidopsis. The location of basic region 1 (BR-1) and 2 (BR-2) is indicated. The leucine residues within the leucine zipper are highlighted and numbered. The hydrophobic amino acids interdigitating the leucine repeats are designated a-d. 219 and 249 denote amino acid positions in GBF-1 asterisks indicate identical amino acids.

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