Biochemical polypeptide chain

The lUBMB Commission on Nomenclature has issued a number of recommendations dealing with areas of a more biochemical nature (72), such as peptide hormones (86), conformation of polypeptide chains (87), abbreviations for nucleic acids and polynucleotides (88), iron—sulfur proteins (89), enzyme units (90), etc. The Commission has also produced rules and recommendations for naming enzymes (91,92). 

Figure 1.2 shows one way of dividing a polypeptide chain, the biochemist s way. There is, however, a different way to divide the main chain into repeating units that is preferable when we want to describe the structural properties of proteins. For this purpose it is more useful to divide the polypeptide chain into peptide units that go from one Ca atom to the next Ca atom (see Figure 1.5). Each C atom, except the first and the last, thus belongs to two such units. The reason for dividing the chain in this way is that all the atoms in such a unit are fixed in a plane with the bond lengths and bond angles very nearly the same in all units in all proteins. Note that the peptide units of the main chain do not involve the different side chains (Figure 1.5). We will use both of these alternative descriptions of polypeptide chains—the biochemical and the structural—and discuss proteins in terms of the sequence of different amino acids and the sequence of planar peptide units.

Many biochemical and biophysical studies of CAP-DNA complexes in solution have demonstrated that CAP induces a sharp bend in DNA upon binding. This was confirmed when the group of Thomas Steitz at Yale University determined the crystal structure of cyclic AMP-DNA complex to 3 A resolution. The CAP molecule comprises two identical polypeptide chains of 209 amino acid residues (Figure 8.24). Each chain is folded into two domains that have separate functions (Figure 8.24b). The larger N-terminal domain binds the allosteric effector molecule, cyclic AMP, and provides all the subunit interactions that form the dimer. The C-terminal domain contains the helix-tum-helix motif that binds DNA. 

Note The formal lUPAC-IUB Commission on Biochemical Nomenclature convention for the definition of the torsion angles polypeptide chain (Biochemistry 9 3471-3479, 1970) is different from that used here, where the atom serves as the point of reference for both rotations, but the result is the same. (Irving Gas)... 

Heliantholysin. The major form of heliantholysin is a basic polypeptide chain (pi in the region of 9.8) having a molecular weight of 16,600. Its amino acid sequence has been determined (11). It is powerfully hemolytic for washed erythrocytes derived from a variety of animals, those of the cat being the most sensitive, and those of the guinea pig the most resistant (10). As is true of most hemolytic systems, the biochemical basis for the very large differences in sensitivity of erythrocytes from different animal species is unknown. 

Most biochemical and biocatalytic studies have been performed with type I B VMOs. This is partly because of the fact that they represent relatively uncomplicated monooxygenase systems. These monooxygenases are typically soluble and composed of only one polypeptide chain. Expression systems have been developed for a number of type I BVMOs while no recombinant expression has been reported for a type II BVMO. Cyclohexanone monooxygenase (CHMO) from an Acinetobacter sp. NCIMB9871 was the only recombinant available BVMO... 

The peptidyl transferase centre of the ribosome is located in the 50S subunit, in a protein-free environment (there is no protein within 15 A of the active site), supporting biochemical evidence that the ribosomal RNA, rather than the ribosomal proteins, plays a key role in the catalysis of peptide bond formation. This confirms that the ribosome is the largest known RNA catalyst (ribozyme) and, to date, the only one with synthetic activity. Adjacent to the peptidyl transferase centre is the entrance to the protein exit tunnel, through which the growing polypeptide chain moves out of the ribosome. 

Tokumoto, T. et al. Regulated interaction between polypeptide chain elongation factor-1 complex with the 26S proteasome during Xenopus oocyte maturation. BMC Biochem 2003, 4, 6. 

Transcription is the term used to describe the synthesis of RNA from a DNA template. Translation is the process by which information in RNA is used to synthesise a polypeptide chain. In a little more detail, the genetic information encoded in DNAis first transcribed into acomplementary copy of RNA (a primary RNA transcript) which is then processed to form messenger RNA (mRNA). This leaves the nucleus and is translated into a polypeptide in the cytosol. This then folds into a three-dimensional structure and may be further biochemically modified (post-transla-tional modification) to produce a protein (Figure 20.18). 

A stretch of DNA that is transcribed as a single continuous RNA strand is called a transcription unit. A unit of transcription may contain one or more sequences encoding different polypeptide chains (translational open reading frames, ORF) or cistrons. The transcription unit is sometimes termed the primary transcript, pre-messenger RNA or heterogeneous nuclear RNA (hnRNA). The primary transcript is further processed to produce mRNA in a form that is relatively stable and readily participates in translation. In order to understand the primary need for processing of this RNA, the biochemical definition of a gene must be discussed. 

This covalent bonding arises as a result of biochemical oxidation of the thiol groups in two cysteine residues, and it may also be achieved chemically with the use of mild oxidizing agents. This modification of thiol groups may thus loop a polypeptide chain or cross-link two separate chains. It also significantly modifies the properties of a... 

AMINO ACIDS, PEPTIDES PROTEINS Recommended nomenclature and symbolism for amino acids and peptides J. Biol Chem. (1985) 260, 14-42 Biochemistry (1975) 14, 449-462 Abbreviations and symbols for the description of the conformation of polypeptide chains /. Biol Chem. (1970) 245, 6489-6497 Nomenclature of iron-sulfur proteins Eur. J. Biochem. (1979) 93, 427-430 Corrections Eur. J. Biochem. (1979) 102, 315 Nomenclature of peptide hormones J. Biol Chem. (1975) 250, 3215-3216 Nomenclature of human immunoglobulins Eur. J. Biochem. (1974) 45, 5-6 Recommended nomenclature of glycoproteins, glyco-peptides, and peptidoglycans /. Biol Chem. (1987) 262, 13-18 Recommended nomenclature of electron-transfer proteins... 

That part of a polypeptide chain(s) possessing catalytic function. The catalytic domain may comprise more than one structural domain and a multienzyme polypeptide contains at least two types of such domains. See Multienzyme Polypeptide Autonomous Catalytic Domain Nomenclature Committee, lUB (1989) Eur. J. Biochem. 185, 485. 

We have shown that out of fifteen forms of three-dimensional crystals from ribosomal particles, grown so far in our laboratory, some appear suitable for crystallographic data collection when using synchrotron radiation at temperatures between 19 °C and —180 °C 50S subunits from H. marismortui., and from B. stearothermophilus, including the -BLl 1 mutant, and the new crystal forms from B. stearothermophilus SOS and Thermus thermophilus 30S subunits which have only recently been grown in non-volatile precipitants We also plan to continue research on biochemically modified particles, such as SOS with one tRNA and its nascent polypeptide chain (which have already been crystallized). 

Frolova LY, Simonsen JL, Merkulova Tl, Litvinov DY, Martensen PM, Rechinsky VO, Camonis JH, Kisselev LL, Justesen J (1998) Functional expression of eukaryotic polypeptide chain release factors 1 and 3 by means of baculovirus/insect cells and complex formation between the factors. Eur J Biochem 256 36 4... 

Veron, M. Falcoz-Kelly E Cohen, G.N. The threonine-sensitive homoserine dehydrogenase and aspartokinase activities of Escherichia coli K12. The two catalytic activities are carried by two independent regions of the polypeptide chain. Eur. J. Biochem., 28, 520-527 (1972)... 

How can a group of compounds, made from a common basis set of amino acids, be so remarkably heterogeneous and exhibit such varied yet specific functions Clearly, the primary structure and the presence or absence of special functional groups, metals, and so on, are of paramount importance. Of complementary importance are the three-dimensional structures of proteins, which are dictated not just by the primary structure but by the way the primary structure is put together biochemically. The polypeptide chains are seldom, if ever, fully extended, but are coiled and folded into more or less stable conformations. As a result, amino-acid side chains in distant positions in the linear sequence are brought into close proximity, and this juxtaposition often is crucial for the protein to fulfill its specific biological function. 

The linear polypeptide chains of a protein fold in a highly specific way that is determined by the sequence of amino acids in the chains. Many proteins are composed of two or more polypeptides. Certain proteins function in structural roles. Some structural proteins interact with lipids in membrane structures. Others aggregate to form part of the cytoskeleton that helps to give the cell its shape. Still others are the chief components of muscle or connective tissue. Enzymes constitute yet another major class of proteins, which function as catalysts that accelerate and direct biochemical reactions. 

The third step is to determine the polypeptide chain end groups. If the polypeptide chains are pure, then only one N-terminal and one C-terminal group should be detected. The amino-terminal amino acid can be identified by reaction with fluorodinitrobenzene (FDNB) (fig. 3.18). Subsequent acid hydrolysis releases a colored dinitrophenol (DNP)-labeled amino-terminal amino acid, which can be identified by its characteristic migration rate on thin-layer chromatography or paper electrophoresis. A more sensitive method of end-group determination involves the use of dan-syl chloride (see Methods of Biochemical Analysis 3B). 

Polypeptide chain end-group analysis, (a) Amino-terminal group identification. A more sensitive method, the dansyl chloride method, is described in Methods of Biochemical Analysis 3B. (b) Carboxyl-terminal group identification. Identification of this amino acid is considerably more difficult. 

Conventionally, the first attribute known about an enzyme used to be its function, usually in a crude extract. This property was screened for in microbial cultures or in tissue samples. The crude extract was then purified to homogeneity and the protein subjected to biochemical studies to learn of its pH and T profiles, its pi and subunit composition, catalytically important residues, and other properties. Proteolytic digestion of the protein with subsequent Edman degradation led to the primary sequence, but no information on the secondary structures such as a-heli-ces and [5-sheets or the folding in three dimensions of the polypeptide chain. The primary sequence could have been used to deduct the gene sequence but, with the degeneration of the code, several possibilities for certain amino acids occur, which makes prediction of the gene sequence a risk. 

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