Molecular architecture of a membrane-spanning hormone acyltransferase required for metabolic regulation

AbstractIntegral membrane proteins represent a large and essential portion of the proteome that often prove challenging for structural studies. We demonstrate a synergistic approach to structurally model topologically complex integral membrane proteins by combining co-evolutionary constraints and computational modeling with biochemical validation. We report the first structural model of a eukaryotic membrane-bound O-acyltransferase (MBOAT), ghrelin O-acyltransferase (GOAT), which modifies the metabolism-regulating hormone ghrelin. Our structure suggests an unanticipated strategy for trans-membrane protein acylation, with catalysis occurring in an internal channel as GOAT acts as an “enzyme inside a pore”. Our structure opens the door to structure-guided inhibitor design targeting GOAT and other MBOAT family members while validating the power of our approach to generate predictive structural models for other experimentally challenging integral membrane proteins.

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Specificity and promiscuity in membrane helix interactions

Quarterly Reviews of Biophysics ◽

10.1017/s0033583500004522 ◽

1994 ◽

Vol 27 (2) ◽

pp. 157-218 ◽

Cited By ~ 127

Author(s):

Mark A. Lemmon ◽

Donald M. Engelman

Keyword(s):

Membrane Proteins ◽

Transmembrane Helix ◽

Integral Membrane Proteins ◽

Lipid Packing ◽

Membrane Protein Folding ◽

Membrane Spanning ◽

Α Helix ◽

Prosthetic Groups ◽

Packing Effects ◽

Helix Interactions

The membrane-spanning portions of many integral membrane proteins consist of one or a number of transmembrane α-helices, which are expected to be independently stable on thermodynamic grounds. Side-by-side interactions between these transmembrane α-helices are important in the folding and assembly of such integral membrane proteins and their complexes. In considering the contribution of these helix–helix interactions to membrane protein folding and oligomerization, a distinction between the energetics and specificity should be recognized. A number of contributions to the energetics of transmembrane helix association within the lipid bilayer will be relatively non-specific, including those resulting from charge–charge interactions and lipid–packing effects. Specificity (and part of the energy) in transmembrane α-helix association, however, appears to rely mainly upon a detailed stereochemical fit between sets of dynamically accessible states of particular helices. In some cases, these interactions are mediated in part by prosthetic groups.

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Membrane-bound annexin V isoforms (CaBP33 and CaBP37) and annexin VI in bovine tissues behave like integral membrane proteins

FEBS Letters ◽

10.1016/0014-5793(92)80369-r ◽

1992 ◽

Vol 296 (2) ◽

pp. 158-162 ◽

Cited By ~ 41

Author(s):

Roberta Bianchi ◽

Ileana Giambanco ◽

Paolo Ceccarelli ◽

Grazia Pula ◽

Rosario Donato

Keyword(s):

Membrane Proteins ◽

Annexin V ◽

Integral Membrane Proteins ◽

Annexin Vi ◽

Membrane Bound

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Membrane Protein Degradation by FtsH Can Be Initiated from Either End

Journal of Bacteriology ◽

10.1128/jb.184.17.4775-4782.2002 ◽

2002 ◽

Vol 184 (17) ◽

pp. 4775-4782 ◽

Cited By ~ 52

Author(s):

Shinobu Chiba ◽

Yoshinori Akiyama ◽

Koreaki Ito

Keyword(s):

Escherichia Coli ◽

Amino Acid ◽

Membrane Proteins ◽

Membrane Protein ◽

Exact Sequence ◽

Integral Membrane Proteins ◽

Amino Acid Residues ◽

Aaa Atpase ◽

Membrane Bound ◽

Periplasmic Domain

ABSTRACT FtsH, a membrane-bound metalloprotease, with cytoplasmic metalloprotease and AAA ATPase domains, degrades both soluble and integral membrane proteins in Escherichia coli. In this paper we investigated how membrane-embedded substrates are recognized by this enzyme. We showed previously that FtsH can initiate processive proteolysis at an N-terminal cytosolic tail of a membrane protein, by recognizing its length (more than 20 amino acid residues) but not exact sequence. Subsequent proteolysis should involve dislocation of the substrates into the cytosol. We now show that this enzyme can also initiate proteolysis at a C-terminal cytosolic tail and that the initiation efficiency depends on the length of the tail. This mode of degradation also appeared to be processive, which can be aborted by a tightly folded periplasmic domain. These results indicate that FtsH can exhibit processivity against membrane-embedded substrates in either the N-to-C or C-to-N direction. Our results also suggest that some membrane proteins receive bidirectional degradation simultaneously. These results raise intriguing questions about the molecular directionality of the dislocation and proteolysis catalyzed by FtsH.

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Ghrelin octanoylation by ghrelin O -acyltransferase: protein acylation impacting metabolic and neuroendocrine signalling

Open Biology ◽

10.1098/rsob.210080 ◽

2021 ◽

Vol 11 (7) ◽

pp. 210080

Author(s):

Tasha R. Davis ◽

Mariah R. Pierce ◽

Sadie X. Novak ◽

James L. Hougland

Keyword(s):

Metabolic Regulation ◽

Energy Homeostasis ◽

Structural Model ◽

Peptide Hormone ◽

Protein Biochemistry ◽

Physiological Processes ◽

Current State ◽

Enzyme Superfamily ◽

Wide Range ◽

Membrane Bound

The acylated peptide hormone ghrelin impacts a wide range of physiological processes but is most well known for controlling hunger and metabolic regulation. Ghrelin requires a unique posttranslational modification, serine octanoylation, to bind and activate signalling through its cognate GHS-R1a receptor. Ghrelin acylation is catalysed by ghrelin O -acyltransferase (GOAT), a member of the membrane-bound O -acyltransferase (MBOAT) enzyme family. The ghrelin/GOAT/GHS-R1a system is defined by multiple unique aspects within both protein biochemistry and endocrinology. Ghrelin serves as the only substrate for GOAT within the human proteome and, among the multiple hormones involved in energy homeostasis and metabolism such as insulin and leptin, acts as the only known hormone in circulation that directly stimulates appetite and hunger signalling. Advances in GOAT enzymology, structural modelling and inhibitor development have revolutionized our understanding of this enzyme and offered new tools for investigating ghrelin signalling at the molecular and organismal levels. In this review, we briefly summarize the current state of knowledge regarding ghrelin signalling and ghrelin/GOAT enzymology, discuss the GOAT structural model in the context of recently reported MBOAT enzyme superfamily member structures, and highlight the growing complement of GOAT inhibitors that offer options for both ghrelin signalling studies and therapeutic applications.

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Adaptation of low-resolution methods for the study of yeast microsomal polytopic membrane proteins: a methodological review

Biochemical Society Transactions ◽

10.1042/bst20120212 ◽

2013 ◽

Vol 41 (1) ◽

pp. 35-42 ◽

Cited By ~ 7

Author(s):

Arlette Bochud ◽

Nagaraju Ramachandra ◽

Andreas Conzelmann

Keyword(s):

Membrane Proteins ◽

Active Sites ◽

Integral Membrane Proteins ◽

The Other ◽

Primary Sequence ◽

Low Resolution ◽

Methodological Review ◽

Binding Domains ◽

Substituted Cysteine Accessibility ◽

Membrane Spanning

Most integral membrane proteins of yeast with two or more membrane-spanning sequences have not yet been crystallized and for many of them the side on which the active sites or ligand-binding domains reside is unknown. Also, bioinformatic topology predictions are not yet fully reliable. However, so-called low-resolution biochemical methods can be used to locate hydrophilic loops or individual residues of polytopic membrane proteins at one or the other side of the membrane. The advantages and limitations of several such methods for topological studies with yeast ER integral membrane proteins are discussed. We also describe new tools that allow us to better control and validate results obtained with SCAM (substituted cysteine accessibility method), an approach that determines the position of individual residues with respect to the membrane plane, whereby only minimal changes in the primary sequence have to be introduced into the protein of interest.

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A high-throughput method for orthophosphate determination of thermostable membrane-bound pyrophosphatase activity

Analytical Methods ◽

10.1039/c7ay02558k ◽

2018 ◽

Vol 10 (6) ◽

pp. 646-651 ◽

Cited By ~ 5

Author(s):

Keni Vidilaseris ◽

Juho Kellosalo ◽

Adrian Goldman

Keyword(s):

Membrane Proteins ◽

Active Transport ◽

High Throughput ◽

Integral Membrane Proteins ◽

Sodium Ions ◽

High Throughput Method ◽

Membrane Bound ◽

Pyrophosphatase Activity

Membrane-bound pyrophosphatases (mPPases) are homodimeric integral membrane proteins that hydrolyse pyrophosphate into orthophosphates coupled to the active transport of protons or sodium ions across membranes.

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Making membrane proteins at the mammalian endoplasmic reticulum

Biochemical Society Transactions ◽

10.1042/bst0311248 ◽

2003 ◽

Vol 31 (6) ◽

pp. 1248-1252 ◽

Cited By ~ 12

Author(s):

F.J.L. Lecomte ◽

N. Ismail ◽

S. High

Keyword(s):

Endoplasmic Reticulum ◽

Protein Synthesis ◽

Membrane Proteins ◽

Lipid Bilayer ◽

Integral Membrane Proteins ◽

Current Understanding ◽

Secretory Proteins ◽

Protein Biogenesis ◽

Membrane Spanning ◽

Membrane Spanning Domains

Whereas protein biogenesis at the endoplasmic reticulum is well understood in the case of secretory proteins and simple membrane proteins, much less is known about the synthesis of multi-spanning integral membrane proteins. While it is clear that the multiple membrane-spanning domains of these proteins must be inserted into the lipid bilayer during biosynthesis, the mechanism by which their integration is achieved and their subsequent folding/assembly are poorly defined. In this review, we summarize our current understanding of protein synthesis at the endoplasmic reticulum and highlight specific features that are relevant to the biogenesis of multi-spanning membrane proteins.

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Image Analysis of Intramembrane Particles in Freeze-Fractured Biomembranes Using Computer Simulation Techniques

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100114694 ◽

1975 ◽

Vol 33 ◽

pp. 214-215

Author(s):

D.J. Benefiel ◽

R.S. Weinstein

Keyword(s):

Membrane Proteins ◽

Freeze Fracture ◽

Integral Membrane Proteins ◽

Systematic Evaluation ◽

Intramembrane Particles ◽

Modeling Methods ◽

Simulation Techniques ◽

Freeze Fracture Electron Microscopy ◽

Fracture Electron ◽

Computer Simulation Techniques

Intramembrane particles (IMP or MAP) are components of most biomembranes. They are visualized by freeze-fracture electron microscopy, and they probably represent replicas of integral membrane proteins. The presence of MAP in biomembranes has been extensively investigated but their detailed ultrastructure has been largely ignored. In this study, we have attempted to lay groundwork for a systematic evaluation of MAP ultrastructure. Using mathematical modeling methods, we have simulated the electron optical appearances of idealized globular proteins as they might be expected to appear in replicas under defined conditions. By comparing these images with the apearances of MAPs in replicas, we have attempted to evaluate dimensional and shape distortions that may be introduced by the freeze-fracture technique and further to deduce the actual shapes of integral membrane proteins from their freezefracture images.

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