Proteins May Prevent Dysfunction and Disease by Relaxing
Determining how proteins function on a molecular level is crucial to understanding the underlying basis
for disease. Now scientists using the U.S. Department of Energy's Advanced Photon Source (APS) at Argonne
National Laboratory are one step closer to unraveling the mystery of how intrinsically disordered proteins
work, according to new research published in Science.
The Chd1 Chromatin Remodeler Shifts Nucleosomal DNA Bidirectionally as a Monomer
Perplexing Cooperative Folding and Stability of a Low-Sequence Complexity, Polyproline 2 Protein Lacking a Hydrophobic Core
DNA in eukaryotic cells is normally found associated with prot4ens called histone in particles called
nucleosomes linked by short segMents of DNA. Chromatin remodelers are specialized ATP-dependent molecular
machines that can reorganize the structure of nucelsomes as needed for such processes such as replication,
transcription and DNA repair. Remodeling results in evenly spaced nucleosomes along the DNA strand yet the
way remodelers achieve this is not understood. CHD-1 is a remodeler important for transcription. Here, the
authors show that the Chd1 remodeler shifts DNA back and forth by dynamically alternating between different
segments of the nucleosome.
The physical basis of protein-folding stability and cooperativity remains a topic of great interest.
Folding of globular proteins is generally assumed to be driven by energetically favorable burial of
hydrophobic groups and that early development of secondary structure increases the cooperativity of
folding. The Sosnick group at the University of Chicago examines these assumptions in a protein (snow
flea antifreeze protein (sfAFP) that is striking in its dearth of hydrophobic burial and its lack of Α
and Β structures, while having a low sequence complexity with 46% glycine. The interior of the protein
is filled only with backbone H-bonds between six polyproline 2 (PP2) helices. Unexpectedly, the protein
folds in a kinetically two-state manner and is moderately stable at room temperature, similar behavior
to that observed for typical globular proteins having Α and Β structures and a hydrophobic core. Hence,
these features are not necessary for folding cooperativity and stability.
Stress-Triggered Phase Separation Is an Adaptive, Evolutionarily Tuned Response
In eukaryotic cells, diverse stresses trigger coalescence of various RNA-binding proteins into
so-called stress granules. In vitro, stress-granule-associated proteins have been observed to demix
to form liquids, hydrogels, and other assemblies. Demixing of an abundant RNA-binding protein into
hydrogel droplets, triggered by stress-associated physiological conditions, appears to promote cell
fitness during stress.
Here, a U Chicago based team lead by D. Allan Drummond and Tobin Sosnick showed that poly(A)-binding
protein (Pab1 in yeast), a defining marker of stress granules, phase separates and forms hydrogels
in-vitro upon exposure to physiological stress conditions.
Asymmetric Unwrapping of Nucleosomal DNA Propagates Asymmetric Opening and Dissociation of the Histone Core
Nucleosomes are protein-DNA structures which eukaryotic organisms use to package and organize DNA
inside the nucleus. Nucleosomes need to be disassembled to permit transcription of DNA and reassembled
afterwards, hence are an important component of the gene regulation machinery. The nucleosome core
particle (NCP) consists of DNA wound around a core of eight histone proteins including two dimers of
H2A-H2B histones and an (H3-H4) tetramer that is assembled as a dimer of dimers. A team led by Lois
Pollack at Cornell University used time-resolved SAXS at the BioCAT beamline 18ID at Advanced Photon
Source and TR-FRET, in collaboration with Lisa Gloss, at Washington State University to study changes
in the DNA conformations as a function of the composition of the histone core during salt induced
disassembly of NCPs in order to allow identification of kinetic pathways and transient intermediates
that show how the sequence of events involving DNA unwrapping and protein dissociation are connected.
The POTRA domains of Toc75 exhibit chaperone-like function to facilitate import into chloroplasts
Chloroplasts, like mitochondria, are organelles of endosymbiotic origin, having evolved from initial
engulfment of a cyanobacterium by a eukaryotic cell. Most of the bacterial genome was subsequently
lost so that most proteins found within chloroplasts are synthesized in the cytoplasm as pre-proteins
and then imported via specialized machinery prior to trafficking to their final destination. Protein
import is accomplished by the TOC (translocon on the outer chloroplast membrane) and TIC (translocon
on the inner chloroplast membrane) machineries in the outer and inner envelope membranes, respectively.
The TOC complex includes a protein called Toc75, which serves as the translocation channel along with
two other proteins, as well as Toc33 and Toc159, which both contain GTPase domains, which help drive
substrate selection and importation. Structural information for the TOC complex was hitherto lacking,
hindering the ability of investigators to form mechanistic models for function. Here a team lead by
Nicholas Noinaj (Purdue University) and Danny Schnell (Michigan State University) reported crystals
structures of Toc75 consisting of three tandem POTRA domains.
Solution Structure of the HIV-1 Intron Splicing Silencer and Its Interactions with the UP1 Domain of Heterogeneous Nuclear Ribonucleoprotein (hnRNP) A1
Human immunodeficiency virus type 1 (HIV-1) requires controlled synthesis of its protein complement
for persistent infection and successful virion production. Genome expression is tightly regulated at
the levels of transcription, splicing, mRNA nuclear export, and translation. RNA polymerase II-dependent
transcription yields a 9-kilobase (kb) polycistronic transcript that undergoes multiple rounds of
alternative splicing to produce upward of 100 different viral mRNAs that recruit antagonistic host
RNA-binding proteins. Thus, HIV-1 splicing pathways are essential components of the viral replication
cycle and represent new targets for therapeutic intervention. Because HIV-1 splicing depends on protein-RNA
interactions, it is important to know the tertiary structures surrounding the splice sites.
Structure of the human elongation factor eEFSec suggest a non-canonical mechanism for selenocysteine incorporation
Selenium is the only essential dietary micronutrient that is genetically encoded in all domains of
life. It is found in proteins as the amino acid selenocysteine (Sec). Mammals, and humans, have 25
ubiquitously expressed selenoproteins, many of which are essential. Selenoproteins and selenoenzymes
are critical for redox potential maintenance, protection of genetic material and cell membrane from
oxidative damage, regulation of the thyroid hormone metabolism, and control of gene expression and
protein folding. This study by the Simonovic group at the University of Illinois at Chicago determined
the crystal structures of the intact human eEFSec in the GTP- and GDP-bound states.
Interaction of TAPBPR, a tapasin homolog, with MHC-I molecules promotes peptide editing
T-cell responses to intracellular pathogens and tumor antigens are governed primarily by effector
CD8+ T cells that recognize antigenic peptides. These peptides are generated by protein degradation
or during translation and are presented at the cell surface by MHC-I molecules. Peptide loading of
major histocompatibility complex class I (MHC-I) molecules is central to antigen presentation,
self-tolerance, and CD8+ T-cell activation. TAP binding protein, related (TAPBPR) is a widely expressed
tapasin homolog that is not part of the classical MHC-I peptide-loading complex (PLC). This report by
the Margulies group, NIH intramural, explores the biochemical and structural basis of the interactions
of TAPBPR with MHC-I molecules.
Unique Bacterial Chemist in the War on Potatoes
In fertile farm soils where potatoes grow, Streptomyces scabies bacteria wage war using chemicals
related to explosives and pesticides. But a microbial spoiler defuses one of S. scabies' poisons.
Researchers at the Georgia Institute of Technology using high-brightness x-rays from the U.S.
Department of Energy's Advanced Photon Source (APS) have gained new insights into a one-of-a-kind
mechanism the microbe employs, which could someday contribute to the development of new agents to
degrade tough pollutants and help rescue crops.
When S. scabies infects potatoes, it spews poisons called thaxtomins, which riddle potatoes with
familiar dark scabs. Perhaps a trifle to the potato connoisseur excising them with a paring knife,
but on a global scale, the blemishes add up to a slash in agricultural production. Scientists
investigating potato soil have found bacteria of the species Bradyrhizobium sp. JS329 running
interference. Though their tough enzymes don't break down thaxtomins, they do render innocuous
another S. scabies toxic secretion called 5-nitroanthranilic acid (5-NAA).
Researchers Find New Clues in 100-Year-Old Mystery of Frank-Starling Law of the Heart
Researchers at Illinois Institute of Technology and Loyola University have discovered new clues in
the 100-year-old mystery of the Frank-Starling law of the heart: What makes the heart contract more
strongly at longer lengths given the same level of calcium activation?
Writing in the February 8 Proceedings of the National Academy of Sciences (PNAS), Biology professor
Thomas Irving of Illinois Tech and Pieter de Tombe of the Loyola University Stritch School of Medicine
demonstrated that the muscle protein titin plays a key role in the Frank-Starling mechanism. The findings
will enable researchers to develop more realistic models of cardiac function and improve their understanding
of cardiac dysfunction in heart failure.
Identifying the Structure of a Tumor-Suppressing Protein
The dimer structure of an important tumor-suppressing protein, phosphatase and tensin homolog
(PTEN) is the second most frequently mutated protein found in human cancer. Phosphatase and
tensin homolog is a known tumor suppressing protein that is encoded by the PTEN gene. When
expressed normally, the protein acts as an enzyme at the cell membrane, instigating a complex
biochemical reaction that regulates the cell cycle and prevents cells from growing or dividing
in an unregulated fashion. Each cell in the body contains two copies of the PTEN gene, one
inherited from each parent. When there is a mutation in one or both of the PTEN genes, it
interferes with the protein's enzymatic activity and, as a result inhibits its tumor suppressing
ability. In order to reveal how dimerization improves PTEN's ability to thwart tumor development.
The researchers in this study, from Carnegie Mellon University, the National Institute of Standards
and Technology, the Center for Synchrotron Radiation Research and Instrumentation, the Illinois
Institute of Technology, Monash University, Harvard Medical School, the University of Massachusetts
Medical School, and Worcester Polytechnic Institute used small-angle x-ray scattering (SAXS) at the
Biophysics Collaborative Access Team 18-ID-D at the APS. Using computer modeling, they found that
in the PTEN dimers, the C-terminal tails of the two proteins may bind the protein bodies in a
cross-wise fashion, which makes them more stable. As a result, they can more efficiently interact
with the cell membrane, regulate cell growth and suppress tumor formation. Now that more is known
about the structure of the PTEN dimer, researchers will be able to use molecular biology tools to
investigate the atomic-scale mechanisms of tumor formation facilitated by PTEN mutations opening up
new avenues for cancer therapeutics.
Microsecond Protein Folding Observed by Time-Resolved SAXS
It is commonly held that random coil polypeptide chains undergo a barrier-less continuous
collapse when the solvent conditions are changed to favor the fully-folded native conformation.
This presumption was tested by probing intramolecular distance distributions during folding of
cytochrome c using time-resolved FRET and continuous turbulent flow mixing SAXS with the
microbeam setup at BioCAT (~80 ms time resolution). Contrary to expectation, tr-FRET revealed
a state with a Trp59-heme distance close to that of the GdnHCl denatured state after ~27 ms of
folding. A concomitant decrease in the population of this state and an increase in the population
of a compact high-FRET state show that the collapse is barrier-limited unlike the barrier less
contraction indicated by single molecule steady state fluorescence studies. SAXS measurements
over a similar time range show that the radius of gyration under native favoring conditions is comparable
to that of the GdnHCl denatured unfolded state. The time courses of SAXS and trFRET were superimposable
and could be fit with two exponential decays. Achieving time resolutions below 100 micro-seconds was
crucial in showing this two component time course.
How a protease may help prevent Alzheimer's disease
Throughout all domains of life, regulated proteolysis ameliorates the effects of
protein damage, misfolding, and aggregation. Unlike canonical protein-protease
networks, M16 metalloproteases, which are Zn2+-dependent and ATP-independent, do
not select substrates on the basis of posttranslational modifications or embedded
degradation tags. These proteases are vital to an array of biological processes,
including the clearance of insulin and other peptide hormones by human insulin-degrading
enzyme (IDE), the removal of targeting peptides from preproteins by the mitochondrial
processing peptidase (MPP, also a component of the cytochrome bc1 complex in plants),
and the catabolism of hemoglobin by falcilysin (Fln) in the malaria parasite).
Human presequence protease (hPreP) is an M16 metalloprotease localized in mitochondria...
These data reveal how hPreP's molecular features enable it to effectively recognize and degrade substrates
in a manner that is flexible, eschewing strict requirements for substrate size, sequence, and
physiochemical profile, yet specific, utilizing defined structural features to destroy amyloidogenic
Fungal prion HET-s as a model for structural complexity and self-propagation in prions
Prions are infectious agents consisting of self-propagating, improperly folded protein
aggregate (amyoloids). Amyloids of the prion protein PrP are responsible for a family of
diseases known as the transmissible spongiform encephalopathies. The fungal prion HET-s
has shown itself to be an excellent model system for studying the inherent properties of
prions. The HET-s prion-forming domain readily folds into a relatively complex two-rung
Β-solenoid amyloid. Investigators at Vanderbilt University used fiber diffraction at
the BioCAT beamline 18ID in combination with site-directed mutagenesis of HET-s to
examine the relative importance of various structural features for the formation of
Β-solenoid structure, as well as study the effects of mutations. Their findings provide
insights into the precise structural interactions necessary for self-propagation of prions.
X-ray Florescence Imaging: A New Tool for Studying Manganese Neurotoxicity
Manganese (Mn) is an essential element required in trace amounts for proper body
function. However, despite its vital role in enzymatic reactions, excessive Mn
exposure leads to a condition known as manganism or Mn induced parkinsonism.
Clinical signs and symptoms of manganism closely resemble those of Parkinson's
disease (PD) and both diseases are pathologically associated with damage to the
basal ganglia. While this condition was first diagnosed about 170 years ago, the
mechanism of the neurotoxic action of Mn remains unknown. Moreover, the possibility
that Mn exposure combined with other genetic and environmental factors can contribute
to the development of Parkinson's disease has been discussed in the literature and
several epidemiological studies have demonstrated a correlation between Mn exposure
and an elevated risk of Parkinson's disease.
NUCLEOTIDE-INDUCED ASYMMETRY WITHIN ATPASE ACTIVATOR RING DRIVES σ54-RNAP
INTERACTION AND ATP HYDROLYSIS
Living creatures use ATP as the "universal energy currency". ATP-ases are assemblies
of molecules that break down ATP into smaller molecules using the energy released
to power myriad biological reactions. Molecular motors are ATP-ases that convert
this chemical energy into mechanical work on other molecules. The AAA+ ATPases are
examples of such molecular machines that perform mechanical work to remodel nearly
every type of macromolecule, in cells from all kingdoms of life. A long-standing,
largely unanswered question about the functional mechanism of the AAA+ ATPases is how
do the rings of chemically identical subunits that make up these assemblies interact
with their target macromolecules?