Structural evidence for dimerization-regulated activation of an integral membrane phospholipase

Snijder, H.J.; Ubarretxena-Belandia, Iban; Blaauw, Mieke; Kalk, K.H.; Verheij, Hubertus M.; Egmond, Maarten R.; Dekker, Niek; Dijkstra, Bauke W.

doi:10.1038/401717a0

Cited by 192 publications

(243 citation statements)

References 29 publications

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“…The different structures are characterized by TM b strands that span the lipid bilayer with an angle of ~45°and often extend into the LPS region of the outer membrane. TM b barrels have an even number of antiparallel TM strands, which is 8 for OmpA [69], 12 for OmPlA [8], 18 for ScrY [11] and 22 for FhuA [12]. OmpA is a small ion channel [29], ScrY is a sucrose-specific porin that forms trimers, FhuA is an active transporter for iron-uptake and OmPlA a phospholipase that forms a dimer.…”

Section: In Vitro Requirements For Membrane Protein Foldingmentioning

confidence: 99%

“…An even number of transmembrane b strands ranging from 8 to 22 has been observed in b-barrel membrane proteins of known crystal structure. Examples are the bacterial outer membrane proteins (OMPs), such as outer membrane protein A (OmpA) with 8 b strands (b-s) [5,6], OmpT (10 b-s) [7], OmPlA (12 b-s) [8], OmpF (16 b-s), maltose (LamB) [9,10] or sucrose (ScrY) [11] porins (18 b-s), and the iron transporters FhuA [12,13] or FepA [14] (22 b-s) (see fig. 1 for some examples).…”

Section: Introductionmentioning

confidence: 99%

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Membrane protein folding on the example of outer membrane protein A of Escherichia coli

Kleinschmidt¹

2003

Cellular and Molecular Life Sciences (CMLS)

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Abstract. The biophysical principles and mechanisms by which membrane proteins insert and fold into a biomembrane have mostly been studied with bacteriorhodopsin and outer membrane protein A (OmpA). This review describes the assembly process of the monomeric outer membrane proteins of Gram-negative bacteria, for which OmpA has served as an example. OmpA is a two-domain outer membrane protein composed of a 171-residue eight-stranded b-barrel transmembrane domain and a 154-residue periplasmic domain. OmpA is translocated in an unstructured form across the cytoplasmic membrane into the periplasm. In the periplasm, unfolded OmpA is kept in solution in complex with the molecular chaperone Skp. After binding of periplasmic lipopolysaccharide, OmpA insertion and folding occur spontaneously upon interaction of the complex with the phospholipid bilayer. Insertion and folding of the b-barrel transmembrane domain into the lipid bilayer are highly synchronized, i.e. the formation of large amounts of b-sheet secondary structure and b-barrel tertiary structure take place in parallel with the same rate constants, while OmpA inserts into the hydrophobic core of the membrane. In vitro, OmpA can successfully fold into a range of model membranes of very different phospholipid compositions, i.e. into bilayers of lipids of different headgroup structures and hydrophobic chain lengths. Three membrane-bound folding intermediates of OmpA were discovered in folding studies with dioleoylphosphatidylcholine bilayers. Their formation was monitored by time-resolved distance determinations by fluorescence quenching, and they were structurally distinguished by the relative positions of the five tryptophan residues of OmpA in projection to the membrane normal. Recent studies indicate a chaperone-assisted, highly synchronized mechanism of secondary and tertiary structure formation upon membrane insertion of b-barrel membrane proteins such as OmpA that involves at least three structurally distinct folding intermediates.

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Section: In Vitro Requirements For Membrane Protein Foldingmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Membrane protein folding on the example of outer membrane protein A of Escherichia coli

Kleinschmidt¹

2003

Cellular and Molecular Life Sciences (CMLS)

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“…Sequence comparisons and evolutionary analyses of lipocalins have led to a classification in which 14 clades have been identified [10]. Bacterial lipocalins belong to Clade 1 (the root clade) while the majority of eukaryotic lipocalins ranging from plants to invertebrates and mammals are found in the higher clades (2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14).…”

Section: Introductionmentioning

confidence: 99%

The membrane bound bacterial lipocalin Blc is a functional dimer with binding preference for lysophospholipids

et al. 2006

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Lipocalins, a widespread multifunctional family of small proteins (15-25 kDa) have been first described in eukaryotes and more recently in Gram-negative bacteria. Bacterial lipocalins belonging to class I are outer membrane lipoproteins, among which Blc from E. coli is the better studied. Blc is expressed under conditions of starvation and high osmolarity, conditions known to exert stress on the cell envelope. The structure of Blc that we have previously solved (V. Campanacci, D. Nurizzo, S. Spinelli, C. Valencia, M. Tegoni, C. Cambillau, FEBS Lett. 562 (2004) 183-188.) suggested its possible role in binding fatty acids or phospholipids. Both physiological and structural data on Blc, therefore, point to a role in storage or transport of lipids necessary for membrane maintenance. In order to further document this hypothesis for Blc function, we have performed binding studies using fluorescence quenching experiments. Our results indicate that dimeric Blc binds fatty acids and phospholipids in a micromolar K d range. The crystal structure of Blc with vaccenic acid, an unsaturated C18 fatty acid, reveals that the binding site spans across the Blc dimer, opposite to its membrane anchored face. An exposed unfilled pocket seemingly suited to bind a polar group attached to the fatty acid prompted us to investigate lyso-phospholipids, which were found to bind in a nanomolar K d range. We discuss these findings in terms of a potential role for Blc in the metabolism of lysophospholipids generated in the bacterial outer membrane.

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“…Crystals suitable for X-ray analysis were grown using detergent solubilised protein and an organic solvent as precipitant [29]. The X-ray structure was published recently [30].…”

Section: Foldmentioning

confidence: 99%

“…2A,B) [30]. Amphipathic L-strands traverse the membrane, presenting a cylindrical hydrophobic protein surface.…”

Section: Foldmentioning

confidence: 99%

Bacterial phospholipase A: structure and function of an integral membrane phospholipase

Snijder

Dijkstra

2000

Biochimica et Biophysica Acta (BBA) - Molecular and Cell Biolog

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Structural evidence for dimerization-regulated activation of an integral membrane phospholipase

Cited by 192 publications

References 29 publications

Membrane protein folding on the example of outer membrane protein A of Escherichia coli

Membrane protein folding on the example of outer membrane protein A of Escherichia coli

The membrane bound bacterial lipocalin Blc is a functional dimer with binding preference for lysophospholipids

Bacterial phospholipase A: structure and function of an integral membrane phospholipase

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