Peroxisome protein import: a complex journey

The import of proteins into peroxisomes possesses many unusual features such as the ability to import folded proteins, and a surprising diversity of targeting signals with differing affinities that can be recognized by the same receptor. As understanding of the structure and function of many components of the protein import machinery has grown, an increasingly complex network of factors affecting each step of the import pathway has emerged. Structural studies have revealed the presence of additional interactions between cargo proteins and the PEX5 receptor that affect import potential, with a subtle network of cargo-induced conformational changes in PEX5 being involved in the import process. Biochemical studies have also indicated an interdependence of receptor–cargo import with release of unloaded receptor from the peroxisome. Here, we provide an update on recent literature concerning mechanisms of protein import into peroxisomes.

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The encapsulin from Thermatoga maritima is a flavoprotein with a symmetry matched ferritin-like cargo protein

10.1101/2021.04.26.441214 ◽

2021 ◽

Author(s):

Benjamin J LaFrance ◽

Caleb Cassidy-Amstutz ◽

Robert J Nichols ◽

Luke M Oltrogge ◽

Eva Nogales ◽

...

Keyword(s):

Enzymatic Activity ◽

Biochemical Analysis ◽

Active Role ◽

Structure And Function ◽

Redox Active ◽

Cargo Protein ◽

Cargo Proteins ◽

And Function ◽

Flavin Binding ◽

Unique Chemical

Bacterial nanocompartments, also known as encapsulins, are an emerging class of protein-based "organelles" found in bacteria and archaea. Encapsulins are virus-like icosahedral particles comprising a ~25-50 nm shell surrounding a specific cargo enzyme. Compartmentalization is thought to create a unique chemical environment to facilitate catalysis and isolate toxic intermediates. Many questions regarding nanocompartment structure-function remain unanswered, including how shell symmetry dictates cargo loading and to what extent the shell facilitates enzymatic activity. Here, we explore these questions using the model T. maritima nanocompartment known to encapsulate a redox-active ferritin-like protein. Biochemical analysis revealed the encapsulin shell to possess a flavin binding site located at the interface between capsomere subunits, suggesting the shell may play a direct and active role in the function of the encapsulated cargo. Furthermore, we used cryoEM to show that cargo proteins use a form of symmetry-matching to facilitate encapsulation and define stoichiometry. In the case of the T. maritima encapsulin, the decameric cargo protein with 5-fold symmetry preferentially binds to the pentameric-axis of the icosahedral shell. Taken together, these observations suggest the shell is not simply a passive barrier-it also plays a significant role in the structure and function of the cargo enzyme.

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The essential activities of the bacterial sigma factor

Canadian Journal of Microbiology ◽

10.1139/cjm-2016-0576 ◽

2017 ◽

Vol 63 (2) ◽

pp. 89-99 ◽

Cited By ~ 16

Author(s):

Maria C. Davis ◽

Christopher A. Kesthely ◽

Emily A. Franklin ◽

Shawn R. MacLellan

Keyword(s):

Gene Expression ◽

Transcription Factors ◽

Rna Synthesis ◽

Gene Expression Regulation ◽

Structure And Function ◽

Structural Studies ◽

General Transcription Factors ◽

Transcription Process ◽

Promoter Dna ◽

And Function

Transcription is the first and most heavily regulated step in gene expression. Sigma (σ) factors are general transcription factors that reversibly bind RNA polymerase (RNAP) and mediate transcription of all genes in bacteria. σ Factors play 3 major roles in the RNA synthesis initiation process: they (i) target RNAP holoenzyme to specific promoters, (ii) melt a region of double-stranded promoter DNA and stabilize it as a single-stranded open complex, and (iii) interact with other DNA-binding transcription factors to contribute complexity to gene expression regulation schemes. Recent structural studies have demonstrated that when σ factors bind promoter DNA, they capture 1 or more nucleotides that are flipped out of the helical DNA stack and this stabilizes the promoter open-complex intermediate that is required for the initiation of RNA synthesis. This review describes the structure and function of the σ70 family of σ proteins and the essential roles they play in the transcription process.

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Advancing an Ecosystem Approach in the Gulf of Maine

Advancing an Ecosystem Approach in the Gulf of Maine ◽

10.47886/9781934874301.ch19 ◽

2012 ◽

Keyword(s):

Trophic Ecology ◽

Gulf Of Maine ◽

Sensor Arrays ◽

Ecosystem Approach ◽

Structure And Function ◽

Ecosystem Dynamics ◽

Dynamical Structure ◽

Factors Affecting ◽

The Future ◽

And Function

<i>Abstract</i> .—Here we summarize presentations given at the theme session “Structure and Function of the Gulf of Maine System” of the 2009 Gulf of Maine Symposium— Advancing Ecosystem Research for the Future of the Gulf, covering a broad spectrum of multidisciplinary research underway in one of the world’s most intensively studied marine systems. Our objective was to attempt a synthesis of the current ecological and oceanographic understanding of the Gulf of Maine and, in particular, to document progress in these areas since the 1996 Gulf of Maine Ecosystem Dynamics Symposium more than a decade earlier. Presentations at the session covered issues ranging from habitat structure and function, biodiversity, population structure, trophic ecology, the intersection of the biological, chemical and physical oceanography of the region, and the dynamics of economically important species. Important strides in characterizing the broader dimensions of biodiversity in the region, the establishment of new sampling programs and the availability of new sensor arrays, and the renewed emphasis synthesis and integration to meet the emerging needs for ecosystem-based management in the gulf have all contributed to a deepened appreciation of its dynamical structure. The critical importance of the ecosystem goods and services provided by the gulf, and the factors affecting the sustainable delivery of these services, was clearly demonstrated in the course of the session. The papers presented at the session made it clear how far we have come and how far we need to go to ensure the sustainable delivery of these services into the future.

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The contribution of voltage clamp fluorometry to the understanding of channel and transporter mechanisms

The Journal of General Physiology ◽

10.1085/jgp.201912372 ◽

2019 ◽

Vol 151 (10) ◽

pp. 1163-1172 ◽

Cited By ~ 8

Author(s):

John Cowgill ◽

Baron Chanda

Keyword(s):

Voltage Clamp ◽

Conformational Changes ◽

Physiological State ◽

Structure And Function ◽

Modern Biology ◽

The Past ◽

Mechanistic Insight ◽

Atomistic Models ◽

Multiple Conformations ◽

And Function

Key advances in single particle cryo-EM methods in the past decade have ushered in a resolution revolution in modern biology. The structures of many ion channels and transporters that were previously recalcitrant to crystallography have now been solved. Yet, despite having atomistic models of many complexes, some in multiple conformations, it has been challenging to glean mechanistic insight from these structures. To some extent this reflects our inability to unambiguously assign a given structure to a particular physiological state. One approach that may allow us to bridge this gap between structure and function is voltage clamp fluorometry (VCF). Using this technique, dynamic conformational changes can be measured while simultaneously monitoring the functional state of the channel or transporter. Many of the important papers that have used VCF to probe the gating mechanisms of channels and transporters have been published in the Journal of General Physiology. In this review, we provide an overview of the development of VCF and discuss some of the key problems that have been addressed using this approach. We end with a brief discussion of the outlook for this technique in the era of high-resolution structures.

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Structural Proteomics Methods to Interrogate the Conformations and Dynamics of Intrinsically Disordered Proteins

Frontiers in Chemistry ◽

10.3389/fchem.2021.603639 ◽

2021 ◽

Vol 9 ◽

Author(s):

Rebecca Beveridge ◽

Antonio N. Calabrese

Keyword(s):

Posttranslational Modifications ◽

Intrinsically Disordered Proteins ◽

Structure And Function ◽

Disordered Proteins ◽

Structural Proteomics ◽

Intrinsically Disordered ◽

Folded Proteins ◽

Structural Mass Spectrometry ◽

And Function ◽

Structural Mass

Intrinsically disordered proteins (IDPs) and regions of intrinsic disorder (IDRs) are abundant in proteomes and are essential for many biological processes. Thus, they are often implicated in disease mechanisms, including neurodegeneration and cancer. The flexible nature of IDPs and IDRs provides many advantages, including (but not limited to) overcoming steric restrictions in binding, facilitating posttranslational modifications, and achieving high binding specificity with low affinity. IDPs adopt a heterogeneous structural ensemble, in contrast to typical folded proteins, making it challenging to interrogate their structure using conventional tools. Structural mass spectrometry (MS) methods are playing an increasingly important role in characterizing the structure and function of IDPs and IDRs, enabled by advances in the design of instrumentation and the development of new workflows, including in native MS, ion mobility MS, top-down MS, hydrogen-deuterium exchange MS, crosslinking MS, and covalent labeling. Here, we describe the advantages of these methods that make them ideal to study IDPs and highlight recent applications where these tools have underpinned new insights into IDP structure and function that would be difficult to elucidate using other methods.

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The encapsulin from Thermotoga maritima is a flavoprotein with a symmetry matched ferritin-like cargo protein

Scientific Reports ◽

10.1038/s41598-021-01932-w ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Benjamin J. LaFrance ◽

Caleb Cassidy-Amstutz ◽

Robert J. Nichols ◽

Luke M. Oltrogge ◽

Eva Nogales ◽

...

Keyword(s):

Biochemical Analysis ◽

Thermotoga Maritima ◽

Active Role ◽

Structure And Function ◽

Redox Active ◽

Cargo Protein ◽

Cargo Proteins ◽

And Function ◽

Flavin Binding ◽

Unique Chemical

AbstractBacterial nanocompartments, also known as encapsulins, are an emerging class of protein-based ‘organelles’ found in bacteria and archaea. Encapsulins are virus-like icosahedral particles comprising a ~ 25–50 nm shell surrounding a specific cargo enzyme. Compartmentalization is thought to create a unique chemical environment to facilitate catalysis and isolate toxic intermediates. Many questions regarding nanocompartment structure–function remain unanswered, including how shell symmetry dictates cargo loading and to what extent the shell facilitates enzymatic activity. Here, we explore these questions using the model Thermotoga maritima nanocompartment known to encapsulate a redox-active ferritin-like protein. Biochemical analysis revealed the encapsulin shell to possess a flavin binding site located at the interface between capsomere subunits, suggesting the shell may play a direct and active role in the function of the encapsulated cargo. Furthermore, we used cryo-EM to show that cargo proteins use a form of symmetry-matching to facilitate encapsulation and define stoichiometry. In the case of the Thermotoga maritima encapsulin, the decameric cargo protein with fivefold symmetry preferentially binds to the pentameric-axis of the icosahedral shell. Taken together, these observations suggest the shell is not simply a passive barrier—it also plays a significant role in the structure and function of the cargo enzyme.

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Nanoscale Analysis of the Effect of Pathogenic Mutations on Polycystin-1

ASME 2010 First Global Congress on NanoEngineering for Medicine and Biology ◽

10.1115/nemb2010-13093 ◽

2010 ◽

Author(s):

Liang Ma ◽

Meixiang Xu ◽

Andres F. Oberhauser

Keyword(s):

Single Molecule ◽

Conformational Changes ◽

Protein Interactions ◽

Structure And Function ◽

Eukaryotic Cells ◽

Mechanical Forces ◽

Force Microscopy ◽

Physiological Conditions ◽

Atomic Force ◽

And Function

The activity of proteins and their complexes often involves the conversion of chemical energy (stored or supplied) into mechanical work through conformational changes. Mechanical forces are also crucial for the regulation of the structure and function of cells and tissues. Thus, the shape of eukaryotic cells is the result of cycles of mechano-sensing, mechano-transduction, and mechano-response. Recently developed single-molecule atomic force microscopy (AFM) techniques can be used to manipulate single molecules, both in real time and under physiological conditions, and are ideally suited to directly quantify the forces involved in both intra- and intermolecular protein interactions. In combination with molecular biology and computer simulations, these techniques have been applied to characterize the unfolding and refolding reactions in a variety of proteins, such as titin (an elastic mechano-sensing protein found in muscle) and polycystin-1 (PC1, a mechanosensor found in the kidney).

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