Halophilic proteins structure

The Genomic Potential Hypothesis holds species to be immutable, and that contention is strongly supported by protein structures. To wit, all members of a species have the same molecules no matter how long they have lived in different parts of the world and members of different species have a larger proportion of different molecules no matter how long they have shared the same living space. This is a strong prediction, true up to the level of the occasional mutation in a species. There are no molecular fossils that could provide clear answers, but a survivor from 250 million years ago, a recently revived halophile,16 was not different from contemporary bacteria ... 

Primary, secondary, tertiary, and quaternary structure are familiar concepts for proteins and refer to the amino acid sequence, local folding arrangement, three-dimensional organization, and subunit interactions of polypeptide chains, respectively. Here, tertiary and quaternary structure shall be considered in the most general way, to include also the small molecules or ions that are essential for the conformational stability of the polypeptide chains. This is especially relevant for halophilic proteins, which have extensive interactions with solvent components (water molecules and salt ions). The known structure of a protein (at any level) always results from experiment, and as such is known only within appropriate error limits. 

Malate dehydrogenase from H. marismortui (AMDH) is the halophilic protein that has been studied most by solution structure methods. A molar mass of 87 kg/mol was determined for the native enzyme. It is stable at high concentrations of NaCl or KC1 and unfolds and dissociates below 2.5 M salt. Pundak and Eisenberg (1981) first measured values for the solvent interactions of AMDH and found that, in contrast to nonhalophilic globular proteins in similar conditions (Bi 0.2—0.3 g/g,B3 0.01 g/g),the halophilic protein bound... 

Halophilic proteins whose solution structures are currently under study include glyceraldehyde-3-phosphate dehydrogenase from Hal-oarcula vallismortis (Krishnan and Altekar, 1990) and a heme-binding catalase-peroxidase from H.marismortui (F. Cendrin, H. Jouvre, and G. Zaccai, private communication). 

It is hoped that development of expression vectors in halobacteria, as well as development of the methodology for gene expression of halophilic proteins in E. coli and their subsequent reactivation, will enable the use of the site-directed mutagenesis methodology in the elucidation of structural features that are responsible for the halophilic properties of these enzymes. 

A section on protein structural chemistry in archaea includes Chapters 5 through 7, respectively, by D. Oesterhelt on the structure and function of photoreceptor proteins in the Halobacteriaceae J. Lanyi on the structure and function of ion-transport rhodopsins in extreme halophiles and R. Hensel on proteins of extreme thermophiles. In a section on cell envelopes (Chapters 8-10), O. Kandler and H. Konig discuss the structure and chemistry of archaeal cell walls M. Kates reviews the chemistry and function of membrane lipids of archaea and L.I. Hochstein covers membrane-bound proteins (enzymes) in archaea. 

In addition to their unusual amino acid content, halophilic proteins need high salt concentrations for maintaining their structure. The volume of halophilic proteins must be measured in these extreme multicomponent solutions. In the case of halophiles, removal of salt would lead to protein denaturation. Some examples for halophilic proteins in concentrated salt solutions are given in Table 11 cf. the values for halophilic malate dehydrogenase and halophilic glutamate dehydrogenase. 

A three-dimensional structure also has been elucidated for bacteriorhodopsin, an integral membrane protein of the halophilic (salt-loving) bacterium Halobacterium halobium. This protein has been studied intensively because of its remarkable activity as a light-driven proton pump (see chapter 14). It forms well-ordered arrays in two-dimensional sheets that can be studied by electron diffraction. Measurements of the diffraction patterns show clearly that bacteriorhodopsin has seven transmembrane helices (fig. 17.12). 

Fig. 7. Schematic representation of AMDH solution structures. The active structures have two parts a catalytically active core, conceivably similar to that in non-halophilic MDH, and protruding loops, required for stabilization in KCl, NaCl, and MgCl2 solvents. In potassium phosphate the protein dimer is stabilized by the hy-drophobicity of the core and the protruding loops are disordered. In KCl (or NaCl) the protein is stabilized by the interaction of the loops in a specific protein—water-salt hydration network. In MgCl2 a similar structure exists with the same amount of water molecules coordinated by fewer salt ions. In low salt concentration, the protein is unfolded and its hydration is like that of nonhalophilic proteins. From Zaccai el al. (1989), with permission.

Oesterhelt, D. (1998). The structure and mechanism of the family of retinal proteins from halophilic archaea. Curr. Opin. Struct. Biol. 8, 489-500. 

Bacteriorhodopsin, is a retinal-containing protein in the purple membrane of a halophilic, (salt-loving) archaebacterium, Halobacterium halobium, which pumps protons out of the cell on activation by light.The three-dimensional structure of bacteriorhodopsin resembles that of rhodopsin in the eye. 

It is clear that more molecular chronometers need to be analysed. On account of the universality of the core metabolic pathways, a molecular study of the enzymes of central metabolism may prove worthwhile in this context. Moreover, the range of phenotypes within the one domain (e.g. extreme halophilicity and thermophilicity, in addition to mesophilicity) may make a comparative study of these enzymes especially valuable to our understanding of the structural basis for extreme protein stability. For these reasons, a number of laboratories are currently engaged in detailed structure-function investigations of the central metabolic enzymes. For a detailed discussion of the comparative enzymology of these pathways, see ref [1]. 

It is now possible to specifically modify the individual rRNA and r-protein molecules that form the archaeal ribosome, enabling one to study the structural/functional relationships of these molecules. Recent studies on the reconstitution of the archaeal 508 ribosomal subunit from the extreme halophile Haloferax mediterranei [20] and the extreme thermophile Sulfolobus solfataricus [21] open the way for the identification of the individual functions of the r-proteins in these particles. Experiments on site-specific changes in the r-protein LI2 from Sulfolobus and their effect on the structure and function of the ribosome are described later in this chapter. 

Yonath, A. High-resolution structures of large ribosomal subunits from mesophilic eubacteria and halophilic archaea at various functional states. Curr. Protein Pept. Sci. 2002, 3(1), 67. 

Fig. 23. (A) The halophilic bacterium H. halobium with patches containing the "purple membrane" (B) Structure of the protein bacteriorhodopsin (left) and the structural formula for the chromophore retinal (right) (C) Covalent binding of retinal with iysine-216 forming a positively-charged Schiff base (D) Illumination of the bacteriorhodopsin retinal and transformation from a trans- to a cis-configuration and releases a proton from the Schiff base to the cell exterior relaxation to ttie trans-form, with uptake of a proton from the cytoplasmic interior. The combination of deprotonation and reprotonation on opposite sides of the membrane constitutes a proton pump. See text for other details. Figures partly adapted from Becker and Deamer (1991) The World of the Cell (2nd ed) Benjamin/Cummings PubI Co. p 215.

Photoactive yellow protein (PYP) was discovered 20 year ago in Halorhodospira halophila, then known as Ectothiorhodospira halophila [1,2]. In several halophilic purple bacteria it has a vital role in the avoidance response to blue light (phototaxis). It has been thoroughly studied as a model photoreceptor system and as the structural prototype for the PAS class of signal transduction proteins. PYP has 125 amino acid residues in an a// -fold with six antiparallel /1-sheets and several helices (see Fig. 5.1). The covalently bound p-coumaric acid chromophore is linked to the only cysteine in the protein (Cys69) (see Fig. 5.1). Hellingwerf has published an excellent review of the photophysical behavior of PYP [1],... 

Bacteriorhodopsin a retinaldehyde-containing purple membrane protein (M, 26,000 248 amino acid residues) first discovered in the halophilic bacterium Halobacierium halobium. The primary structure of B. is not homologous with that of vertebrate rhodopsin (M, 40,000), but the tertiary structures of the two proteins are similar. B. consists of 7 a-helical regions which lie in the membrane. These are connected by nonhelical regions which protrude into the cytoplasm and extracellular space. The membrane portions of the mole-... 

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