Multimeric molecules

Protein molecules that have only one chain are called monomeric proteins. But a fairly large number of proteins have a quaternary structure, which consists of several identical polypeptide chains (subunits) that associate into a multimeric molecule in a specific way. These subunits can function either independently of each other or cooperatively so that the function of one subunit is dependent on the functional state of other subunits. Other protein molecules are assembled from several different subunits with different functions for example, RNA polymerase from E. coli contains five different polypeptide chains. 

Some proteins are formed by a single chain and are called monomeric, but a large number are formed by several polypeptide chains that associate in a multimeric molecule. The relationships of the peptide chains in a multichain protein are known as the quaternary structure. These subunits may work either independently of each other or cooperatively, i.e. the function of one subunit depends on the functional state of the others11. 

The terms polypeptide and protein are used interchangeably in discussing single polypeptide chains. The term protein broadly defines molecules composed of one or more polypeptide chains. Proteins having only one polypeptide chain are monomeric proteins. Proteins composed of more than one polypeptide chain are multimeric proteins. Multimeric proteins may contain only one kind of polypeptide, in which case they are homomultimeric, or they may be composed of several different kinds of polypeptide chains, in which instance they are heteromultimeric. Greek letters and subscripts are used to denote the polypeptide composition of multimeric proteins. Thus, an ag type protein is a dimer of identical polypeptide subunits, or a homodimer. Hemoglobin (Table 5.1) consists of four polypeptides of two different kinds it is an hetero-multimer. 

If the protein of interest is a heteromultimer (composed of more than one type of polypeptide chain), then the protein must be dissociated and its component polypeptide subunits must be separated from one another and sequenced individually. Subunit associations in multimeric proteins are typically maintained solely by noncovalent forces, and therefore most multimeric proteins can usually be dissociated by exposure to pEI extremes, 8 M urea, 6 M guanidinium hydrochloride, or high salt concentrations. (All of these treatments disrupt polar interactions such as hydrogen bonds both within the protein molecule and between the protein and the aqueous solvent.) Once dissociated, the individual polypeptides can be isolated from one another on the basis of differences in size and/or charge. Occasionally, heteromultimers are linked together by interchain S—S bridges. In such instances, these cross-links must be cleaved prior to dissociation and isolation of the individual chains. The methods described under step 2 are applicable for this purpose. 

All of the transport systems examined thus far are relatively large proteins. Several small molecule toxins produced by microorganisms facilitate ion transport across membranes. Due to their relative simplicity, these molecules, the lonophore antibiotics, represent paradigms of the mobile carrier and pore or charmel models for membrane transport. Mobile carriers are molecules that form complexes with particular ions and diffuse freely across a lipid membrane (Figure 10.38). Pores or channels, on the other hand, adopt a fixed orientation in a membrane, creating a hole that permits the transmembrane movement of ions. These pores or channels may be formed from monomeric or (more often) multimeric structures in the membrane. 

Hemoglobins bind four molecules of Oj per tetramer, one per heme. A molecule of Oj binds to a hemoglobin tetramer more readily if other Oj molecules are already bound (Figure 6-4). Termed cooperative binding, this phenomenon permits hemoglobin to maximize both the quantity of O2 loaded at the PO2 of the lungs and the quantity of O2 released at the PO2 of the peripheral tissues. Gooperative interactions, an exclusive property of multimeric proteins, are critically important to aerobic life. 

Because mechanism-based inactivation depends on enzyme catalysis, there cannot be more than one molecule of inactivator bound to the enzyme active site. Thus formation of the covalent E-A species cannot result in a stoichiometry of inactivator to enzyme of greater than 1 1. In the case of multimeric enzymes, however, it may not be necessary to covalently modify all of the enzyme active sites within the multi-mer in order to effect total inactivation of the enzyme. In this situation one may observe a stoichiometry of less that 1 1. Under no circumstances, however, can a mechanism-based inactivator display a stoichiometry of greater than 1 1 with the enzyme. 

LDMS is particularly well suited for the analysis of porphyrins.35-39 The heme molecule—a 22 rc-electron conjugated protoporphyrin system (Figure 8.1)—is an efficient photo-absorber in the visible and near UV (with an absorption maximum—the Soret band—near 400nm). This feature, concurrently with its low ionization potential, warrants that direct LDMS will possess extremely low limits for heme detection. The uses of IR or UV LDMS for structural characterization of natural porphyrins and their metabolites, synthetic monomeric porphyrins (e.g., used in photodynamic therapy), porphyrin polymers, and multimeric arrays, have been well documented.41148 In addition fast atom bombardment MS has been used to characterize purified hemozoin, isolated from the spleens and livers of Plasmodium yoelii infected mice.49... 

The general types of protein-protein interactions that occur in cells include receptor-ligand, enzyme-substrate, multimeric complex formations, structural scaffolds, and chaperones. However, proteins interact with more targets than just other proteins. Protein interactions can include protein-protein or protein-peptide, protein-DNA/RNA or protein-nucleic acid, protein-glycan or protein-carbohydrate, protein-lipid or protein-membrane, and protein-small molecule or protein-ligand. It is likely that every molecule within a cell has some kind of specific interaction with a protein. 

In the tight of the results presented above, we conclude that alfalfa offers a suitable system for the high-yield production of correctly assembled complex proteins, including multimeric glycoproteins. The post-translational capacities of alfalfa indicate that this system is one of the best-suited for the production of molecules for therapeutic and diagnostic applications. 

Fujita and coworkers have also reported the encapsulation of multimeric porphyrin assemblies in the box-shaped cavities of ternary Pd6 coordination cages. Two types of cofacial porphine dimers A and B could be stabilized (133). In the smaller [Pd6(L14)2(L15)6]12+ cage 29, whose diameter is 10.4 A (Fig. 21), two porphyrin molecules can be stackedo directly on top of each other with an interplane distance of 3.4 A. In the larger cage 30, an additional molecule of L14 is intercalated between the two porphyrin bases. All complexes were found to be water-soluble in contrast to other 7i-stacked porphyrin dimers. The encapsulation... 

Carpenter et al. [1.120] found that certain polymers (e. g. PVP) could stabilize multimeric enzymes during freezing and freeze drying by a different mechanism They cannot replace water molecules in the dried state therefore it is assumed that they inhibited the dissociation of the enzymes molecules induced by freezing and freeze drying. 

In addition to the binding of substrate (or in some cases co-substrates) at the active site, many enzymes have the capacity to bind regulatory molecules at sites which are usually spatially far removed from the catalytic site. In fact, allosteric enzymes are invariably multimeric (i.e. have a quaternary structure) and the allosteric (regulatory) sites are on different subunits of the protein to the active site. In all cases, the binding of the regulatory molecules is non covalent and is described in kinetic terms as noncompetitive inhibition. 

Each of these proteins is blue and appears to have a minimum of four copper atoms per molecule one type 1, one type 11, and two type III. Laccase is not known to be multimeric, nor is ceruloplasmin, but ascorbate oxidase is apparently a dimer. 

---