514 publications from this institution
Membrane proteins present a hydrophobic surface to the surrounding lipid, whereas portions protruding into the aqueous milieu expose a polar surface. But how have proteins evolved to deal with the complex environment at the membrane–water interface? Some insights have been provided by high-resolution structures of membrane proteins, and recent studies of the role of individual amino acids in mediating protein–lipid contacts have shed further light on this issue. It now appears clear that the polar-aromatic residues Trp and Tyr have a specific affinity for a region near the lipid carbonyls, whereas positively charged residues extend into the lipid phosphate region.
Thanks to the rapid progress in membrane protein crystallography and cryogenic electron microscopy in recent years, we have become increasingly accustomed to seeing protein structures characterized by long transmembrane α-helices (TMHs) crisscrossing a lipid bilayer (1). But how is it decided which segments of the polypeptide form the TMHs, and what are the precise sequence requirements for their membrane insertion? The great majority of integral membrane proteins require one or another type of translocon to catalyze insertion into the target membrane. The most commonly used and best understood of these are the universally conserved Sec-type translocons found in the cytoplasmic membrane of bacteria (the SecYEG complex) and the endoplasmic reticulum (ER) membrane in eukaryotes (the Sec61 complex) (2). They mediate cotranslational membrane insertion of the growing polypeptide as it emerges from the exit tunnel of translocon-bound ribosomes (3), and have an evolutionary conserved structure with a central channel through which hydrophilic polypeptide segments can be translocated across the membrane and a dynamic “lateral gate” through which hydrophobic segments in the nascent polypeptide chain can partition into the surrounding lipid bilayer to form TMHs (4, 5). Over the past two decades, statistical and experimental studies have progressively uncovered the sequence characteristics that define the difference between polypeptide segments that can form TMHs and those that cannot. To a first approximation, the efficiency of membrane insertion of a TMH is dictated by its hydrophobicity, although interactions with neighboring TMHs, or sequence variations in segments immediately flanking the TMH, can shift the hydrophobicity threshold (HT) required for … [↵][1]1Email: gunnar{at}dbb.su.se. [1]: #xref-corresp-1-1
The bacterial Sec genes encode a generalized protein export machinery. Although the mitochondria present in eukaryotic cells are derived from bacterial ancestors, a comprehensive search of the complete genomic sequence for the eukaryotic yeast Saccharomyces cerevisiae did not reveal any close homologs of the bacterial Sec genes, strongly suggesting that yeast mitochondria lack a generalized bacterial‐type export system. This finding has implications for the sorting of imported mitochondrial proteins to the intermembrane space compartment, and also for the insertion of mitochondrially encoded proteins into the inner membrane.
Integral membrane proteins from a wide variety of sources conform to a positive-inside rule, with many more positively charged amino acids in their cytoplasmic as compared to extracytoplasmic domains. A growing body of experimental work also points to positively charged residues in regions flanking the apolar transmembrane segments as being the main topological determinants. In this paper, we report a systematic comparison of the effects of positively (Arg, Lys, His) as well as negatively (Asp, Glu) charged residues on the membrane topology of a model Escherichia coli inner membrane protein. Our results show that positive charge is indeed the major factor determining the transmembrane topology, with Arg and Lys being of nearly equal efficiency. His, although normally a very weak topological determinant, can be potentiated by a lowering of the cytoplasmic pH. Asp and Glu affect the topology to similar extents and only when present in very high numbers.
No abstract is provided for this article.
We show that the so‐called ‘positive inside’ rule, i.e. the observation that positively charged amino acids tend to be more prevalent in cytoplasmic than in extra‐cytoplasmic segments in transmembrane proteins [von Heijne, G. (1986) EMBO J. 5 , 3021–3027], seems to hold for all polar segments in multi‐spanning eukaryotic membrane proteins irrespective of their position in the sequence and hence can be used in conjunction with hydrophobicity analysis to predict their transmembrane topology. Further, as suggested by others, we confirm that the net charge difference across the first transmembrane segment correlates well with its orientation [Hartmann, E., Rapoport, T. A. and Lodish, H. F. (1989) Proc. Natl Acad. Sci. USA 86 , 5786–5790], and that the overall amino‐acid composition of long polar segments can also be used to prodict their cytoplasmic or extra‐cytoplasmic location [Nakashima, H. and Nishikawa, K. (1992) FEBS Lett. 303 , 141–146]. We present an approach to the topology prediction problem for eukaryotic membrane proteins based on a combination of these methods.
We have mapped the topology of the C-terminal half of the putative potassium channel protein Kch in the inner membrane of Escherichia coli using PhoA fusions. Our results are consistent with the widely assumed six-transmembrane helix model for eukaryotic voltage-gated ion channels and place both ends of the proposed channel-lining P-segment on the periplasmic side of the inner membrane. The rather hydrophobic P-segment is found to translocate only slowly across the inner membrane and seems to be near a threshold for stop-transfer function. We have mapped the topology of the C-terminal half of the putative potassium channel protein Kch in the inner membrane of Escherichia coli using PhoA fusions. Our results are consistent with the widely assumed six-transmembrane helix model for eukaryotic voltage-gated ion channels and place both ends of the proposed channel-lining P-segment on the periplasmic side of the inner membrane. The rather hydrophobic P-segment is found to translocate only slowly across the inner membrane and seems to be near a threshold for stop-transfer function.
Two-component signaling circuits allow bacteria to detect and respond to external stimuli. Unfortunately, the input stimulus remains unidentified for the majority of these circuits. Therefore, development of a synthetic method for stimulus-independent modulation of these circuits is highly desirable because particular physiological or developmental processes could be controlled for biotechnological purposes without the need to identify the stimulus itself. Here, we demonstrate that aromatic tuning, i.e., repositioning the aromatic residues commonly found at the cytoplasmic end of the receptor (EnvZ) transmembrane domain, facilitates stimulus-independent modulation of signal output from the EnvZ/OmpR osmosensing circuit of Escherichia coli. We found that these osmosensing circuits retained the ability to respond appropriately to increased external osmolarity, suggesting that the tuned receptors were not locked in a single conformation. We also noted that circuits containing aromatically tuned variants became more sensitive to changes in the receptor concentration than their wild-type counterpart, suggesting a new way to study mechanisms underpinning receptor concentration-dependent robustness. We believe that aromatic tuning has several advantages compared to previous methods aimed at stimulus-independent modulation of receptors and that it will be generally applicable to a wide-range of two-component circuits.
ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTThree-dimensional model for the membrane domain of Escherichia coli leader peptidase based on disulfide mappingPaul Whitley, Lennart Nilsson, and Gunnar von HeijneCite this: Biochemistry 1993, 32, 33, 8534–8539Publication Date (Print):August 24, 1993Publication History Published online1 May 2002Published inissue 24 August 1993https://pubs.acs.org/doi/10.1021/bi00084a020https://doi.org/10.1021/bi00084a020research-articleACS PublicationsRequest reuse permissionsArticle Views98Altmetric-Citations43LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InRedditEmail Other access optionsGet e-Alertsclose Get e-Alerts
No abstract is provided for this article.
The SecY (bacteria) and Sec61 (eukaryotes) translocon complexes, or protein-conducting channels, work in concert with bound ribosomes to insert proteins into membranes during the first step of membrane protein assembly. The crystallographic structure of an archaeal SecY translocon provides dramatic new insights into the mechanism of translocon function. This structure suggests an explanation for how the translocon can aid in establishing membrane protein topology via the positive-inside rule. The folding of membrane proteins may begin in the ribosome exit tunnel, before entering the translocon, according to cryo-electron microscopy and biophysical studies.
