Gel-like Aggregates Research Articles

Biomolecular Condensates: Structure, Functions, Methods of Research.

The term "biomolecular condensates" is used to describe membraneless compartments in eukaryotic cells, accumulating proteins and nucleic acids. Biomolecular condensates are formed as a result of liquid-liquid phase separation (LLPS). Often, they demonstrate properties of liquid-like droplets or gel-like aggregates; however, some of them may appear to have a more complex structure and high-order organization. Membraneless microcompartments are involved in diverse processes both in cytoplasm and in nucleus, among them ribosome biogenesis, regulation of gene expression, cell signaling, and stress response. Condensates properties and structure could be highly dynamic and are affected by various internal and external factors, e.g., concentration and interactions of components, solution temperature, pH, osmolarity, etc. In this review, we discuss variety of biomolecular condensates and their functions in live cells, describe their structure variants, highlight domain and primary sequence organization of the constituent proteins and nucleic acids. Finally, we describe current advances in methods that characterize structure, properties, morphology, and dynamics of biomolecular condensates in vitro and in vivo.

Scale-Dependent Rheology of Synovial Fluid Lubricating Macromolecules.

Synovial fluid (SF) is the complex biofluid that facilitates the exceptional lubrication of articular cartilage in joints. Its primary lubricating macromolecules, the linear polysaccharide hyaluronic acid (HA) and the mucin-like glycoprotein proteoglycan 4 (PRG4 or lubricin), interact synergistically to reduce boundary friction. However, the precise manner in which these molecules influence the rheological properties of SF remains unclear. This study aimed to elucidate this by employing confocal microscopy and multiscale rheometry to examine the microstructure and rheology of solutions containing recombinant human PRG4 (rhPRG4) and HA. Contrary to previous assumptions of an extensive HA-rhPRG4 network, it is discovered that rhPRG4 primarily forms stiff, gel-like aggregates. The properties of these aggregates, including their size and stiffness, are found to be influenced by the viscoelastic characteristics of the surrounding HA matrix. Consequently, the rheology of this system is not governed by a single length scale, but instead responds as a disordered, hierarchical network with solid-like rhPRG4 aggregates distributed throughout the continuous HA phase. These findings provide new insights into the biomechanical function of PRG4 in cartilage lubrication and may have implications in the development of HA-based therapies for joint diseases likeosteoarthritis.

M1A in CAG repeat RNA binds to TDP-43 and induces neurodegeneration

Microsatellite repeat expansions within genes contribute to a number of neurological diseases1,2. The accumulation of toxic proteins and RNA molecules with repetitive sequences, and/or sequestration of RNA-binding proteins by RNA molecules containing expanded repeats are thought to be important contributors to disease aetiology3–9. Here we reveal that the adenosine in CAG repeat RNA can be methylated to N1-methyladenosine (m1A) by TRMT61A, and that m1A can be demethylated by ALKBH3. We also observed that the m1A/adenosine ratio in CAG repeat RNA increases with repeat length, which is attributed to diminished expression of ALKBH3 elicited by the repeat RNA. Additionally, TDP-43 binds directly and strongly with m1A in RNA, which stimulates the cytoplasmic mis-localization and formation of gel-like aggregates of TDP-43, resembling the observations made for the protein in neurological diseases. Moreover, m1A in CAG repeat RNA contributes to CAG repeat expansion-induced neurodegeneration in Caenorhabditis elegans and Drosophila. In sum, our study offers a new paradigm of the mechanism through which nucleotide repeat expansion contributes to neurological diseases and reveals a novel pathological function of m1A in RNA. These findings may provide an important mechanistic basis for therapeutic intervention in neurodegenerative diseases emanating from CAG repeat expansion.

Charge Segregation in the Intrinsically Disordered Region Governs VRN1 and DNA Liquid-like Phase Separation Robustness

VERNALIZATION1 (VRN1) is a transcriptional repressor involved in plant vernalization that undergoes liquid–liquid phase separation (LLPS) with DNA. The naturally occurring VRN1-like proteins contain two B3 DNA binding domains connected by an intrinsically disordered region (IDR). The IDR length in VRN1-like proteins has a broad distribution, while the charge segregation pattern is largely conserved. We studied the effect of IDR length and charge segregation on DNA-induced VRN1 phase separation. When only neutral residues (Pro-Ser repeats) were used, the phase separation behavior is sensitive to IDR length, changing from gel-like aggregates (L = 40) to liquid-like droplets (L = 100–120) and clear solution (L = 160). When a pair of continuous patches of positive and negative residues were added to the IDRs, all the VRN1 variants formed robust and durable droplets with DNA independent of the IDR length. To test how robust the system is, we introduced folded green fluorescent protein or the enzyme GPX4 into VRN1 variants with charge segregation in IDR, the resulting proteins form LLPS with DNA as well. Our study implies that VRN1-like proteins use conserved charge segregation pattern to retain functional LLPS during evolution, and demonstrates the possibility of using this system to design novel biosensors or bio-factories by introducing various functional modules.

Nanosized Phase Segregation of Sphingomyelin and Dihydrosphigomyelin in Unsaturated Phosphatidylcholine Binary Membranes without Cholesterol.

In this study, we applied fluorescence spectroscopy, differential scanning calorimetry (DSC), and 2H NMR to elucidate the properties of nanoscopic segregated domains in stearoylsphingomyelin (SSM)/dioleoylphosphatidylcholine (DOPC) and dihydrostearoylsphingomyelin (dhSSM)/DOPC binary membranes. The results obtained from fluorescence measurements suggest the existence of gel-like domains with high fluidity in both SSM and dhSSM macroscopic gel phases. The DSC thermograms showed that DOPC destabilizes SM-rich gel-like domains to a much lesser extent compared to the same amount of cholesterol. It was also found that a stable lateral segregation occurs without cholesterol, indicating that SSM itself undergoes homophilic interactions to form small gel-like domains. 2H NMR experiments disclosed differences in the temperature-dependent ordering of SSM/DOPC and dhSSM/DOPC bilayers; the dhSSM membrane showed less miscibility with the DOPC fluid phase, higher thermal stability, and tighter packing. In addition, the NMR results suggest the formation of mid-sized gel-like aggregates consisting of dhSSM. These differences could be accounted for by homophilic interactions, as previously reported ( Yasuda Biophys. J. 2016 , 110 , 431 - 440 ). In the absence of cholesterol, the moderately strong sphingomyelin (SM)/SM affinity results in the formation of small gel-like domains, whereas a stronger dhSSM/dhSSM affinity leads to larger gel-like domains. Considering the similar physicochemical features of SSM and dhSSM, the present results suggest that the formation of nanosized domains of SM is better characterized by homophilic interactions than by SM-cholesterol interplay. These effects are considered important to the ordered domain formation of SMs in biological membranes.

Unusual Aggregation Arrangement of Eu-Containing Polyoxometalate Hybrid in a Protic Ionic Liquid with Improved Luminescence Property.

Hybridization of polyoxometalates (POMs) with cationic surfactants offers the opportunity to greatly improve their functionalities as well as processabilities. Here, a surfactant-encapsulated Eu-containing POM complex (SEP) was formed via electrostatic interaction between 1-octadecyl-3-methylimidazolium bromide (OB) and Na9(EuW10O36)·32H2O (EuW10). SEP was first self-assembled in a protic ionic liquid to prepare the soft aggregates to fundamentally avoid the fluorescence quenching by water molecules. The structures and photophysical properties of SEP or aggregates were investigated thoroughly by NMR and FTIR spectroscopy, optical and electron microscopy, small-angle X-ray scattering, and fluorescence measurements. The formed gel-like aggregates were found to compose of three-dimensional networks of microribbons with an interdigitated layered molecular packing of SEP, which was different from the usual inverse bilayer model of POM hybrids in common organic solvents. Compared to EuW10 solid or its aqueous solution, both SEP and its aggregates exhibited intense red luminescence with much improved lifetime and quantum efficiency. In addition, the soft aggregates exhibited an efficient energy transfer and an obviously enhanced monochromaticity, owning to the organized arrangement of EuW10 units and a confined microenvironment to isolate them from each other between adjacent layers. The obtained results will not only present a useful reference to the aggregation behavior of POM hybrids in ionic liquids, but also provide an easy way to design EuW10 luminescent soft materials based on the nonaqueous media.

Different characteristic effects of ageing on starch-based films plasticised by 1-ethyl-3-methylimidazolium acetate and by glycerol

The focus of this study was on the effects of plasticisers (the ionic liquid 1-ethyl-3-methylimidazolium acetate, or [Emim][OAc]; and glycerol) on the changes of starch structure on multiple length scales, and the variation in properties of plasticised starch-based films, during ageing. The films were prepared by a simple melt compression moulding process, followed by storage at different relative humidity (RH) environments. Compared with glycerol, [Emim][OAc] could result in greater homogeneity in [Emim][OAc]-plasticised starch-based films (no gel-like aggregates and less molecular order (crystallites) on the nano-scale). Besides, much weaker starch–starch interactions but stronger starch-[Emim][OAc] interactions at the molecular level led to reduced strength and stiffness but increased flexibility of the films. More importantly, [Emim][OAc] (especially at high content) was revealed to more effectively maintain the plasticised state during ageing than glycerol: the densification (especially in the amorphous regions) was suppressed; and the structural characteristics especially on the nano-scale were stabilised (especially at a high RH), presumably due to the suppressed starch molecular interactions by [Emim][OAc] as confirmed by Raman spectroscopy. Such behaviour contributed to stabilised mechanical properties. Nonetheless, the crystallinity and thermal stability of starch-based films with both plasticisers were much less affected by ageing and moisture uptake during storage (42 days), but mostly depended on the plasticiser type and content. As starch is a typical semi-crystalline bio-polymer containing abundant hydroxyl groups and strong hydrogen bonding, the findings here could also be significant in creating materials from other similar biopolymers with tailored sensitivity and properties to the environment.

Nonionic cyclodextrin based binary system with upper and lower critical solution temperature transitions via supramolecular inclusion interaction.

A nonionic binary aqueous interaction system consisting of β-cyclodextrin trimer (β-CD3) and naphthalene-terminated poly(ethylene glycol) (PEG-NP2), which has tunable upper critical solution temperature (UCST) behavior around room temperature and lower critical solution temperature (LCST) behavior at high temperature, was investigated. In the UCST transition, gel-like aggregates form because of supramolecular inclusion complexation between β-CD3 and PEG-NP2. During LCST transition, PEG-NP2 becomes insoluble in water, which results in its precipitation. The effects of concentration, stoichiometry of the two components, and electrolyte on UCST behavior are discussed. This study provides a new nonionic thermoresponsive material.

Liquid-crystal mediated nanoparticle interactions and gel formation

Colloidal particles embedded within nematic liquid crystals exhibit strong anisotropic interactions arising from preferential orientation of nematogens near the particle surface. Such interactions are conducive to forming branched, gel-like aggregates. Anchoring effects also induce interactions between colloids dispersed in the isotropic liquid phase, through the interactions of the pre-nematic wetting layers. Here we utilize computer simulation using coarse-grained mesogens to perform a molecular-level calculation of the potential of mean force between two embedded nanoparticles as a function of anchoring for a set of solvent conditions straddling the isotropic-nematic transition. We observe that strong, nontrivial interactions can be induced between particles dispersed in mesogenic solvent, and explore how such interactions might be utilized to induce a gel state in the isotropic and nematic phases.

ESR studies of nitrogen atoms stabilized in aggregates of krypton–nitrogen nanoclusters immersed in superfluid helium

Impurity–helium condensates (IHCs) containing nitrogen and krypton atoms immersed in superfluid 4He have been studied via CW electron spin resonance (ESR). The IHCs are gel-like aggregates of nanoclusters composed of impurity species. It was found that the addition of krypton atoms to the nitrogen–helium gas mixture used for preparation of IHCs increases the efficiency of stabilization of nitrogen atoms. We have achieved high average (5 · 1019 cm−3) and local (2 · 1021 cm−3) concentrations of nitrogen atoms in krypton–nitrogen–helium condensates. The analysis of ESR lines shows that in krypton-nitrogen nanoclusters three different sites exist for stabilization of nitrogen atoms. Nitrogen atoms are stabilized in the krypton core of nanoclusters, in the nitrogen molecular layer that covers the Kr core, and on the surface of the nanoclusters. High concentrations of nitrogen atoms achieved in IHCs provide an important step in the search for magnetic ordering effects at low temperatures.

Thermally induced transformations of calcium carbonate polymorphs precipitated selectively in ethanol/water solutions

For the precipitation of calcium carbonate polymorphs in ethanol/water solutions of calcium chloride by the diffusion of the gases produced by sublimation–decomposition of solid ammonium carbonate, polymorph selection and morphology control of the precipitates were demonstrated by the effect of ethanol/water ratio in the mother liquor. The precipitated phases change systematically from gel-like aggregates of hydrated amorphous calcium carbonate in the absolute ethanol solution to well-shaped rhombohedral particles of calcite in the absolute aqueous solution via almost pure phase of vaterite with dendrite structure in 75%-ethanol/25%-aqueous and 50%-ethanol/50%-aqueous solutions. On heating the precipitated sample in flowing dry nitrogen, all the samples transformed to calcite before the thermal decomposition, where the thermal decomposition temperature shifts to higher temperatures with increasing the water content in the mother liquor due to the systematic increase in the particle size of calcite. Accordingly, the present method of controlled precipitation of calcium carbonate polymorphs is also useful to control the particle size and reactivity of calcite produced by heating the precipitates. Selecting vaterite with dendrite structure from the present series of precipitated samples, the structural phase transition to calcite was characterized as the three-dimensional growth of rhombohedral particles of calcite with the enthalpy change Δ H = − 2.8 ± 0.1 kJ mol −1 and the apparent activation energy E a = 289.9 ± 5.8 kJ mol −1.

Stabilization of hydrogen atoms in aggregates of krypton nanoclusters immersed in superfluid helium

Impurity-helium condensates (IHCs) containing krypton and hydrogen atoms immersed in superfluid $^{4}\text{H}\text{e}$ have been studied via cw electron-spin-resonance (ESR) techniques. The IHCs are gel-like aggregates of nanoclusters composed of krypton and hydrogen atoms. We have found that very high average $(\ensuremath{\sim}{10}^{19}\text{ }{\text{cm}}^{\ensuremath{-}3})$ and local $(\ensuremath{\sim}6\ifmmode\times\else\texttimes\fi{}{10}^{19}\text{ }{\text{cm}}^{\ensuremath{-}3})$ concentrations of hydrogen atoms can be stabilized in these samples. The analysis of ESR line positions and shapes shows that most of the H atom population resides in solid ${\text{H}}_{2}$ layers on the krypton nanocluster surfaces. High concentrations of atomic hydrogen achieved in IHCs provide an important step in the search for collective magnetic and quantum statistical phenomena at lower temperatures.

CW ESR studies of impurity-helium condensates containing krypton and hydrogen atoms

Impurity-Helium condensates (IHCs) containing krypton and hydrogen atoms have been studied in superfluid helium-4 via CW ESR techniques. The IHCs studied in this work are gel-like aggregates of nanoclusters composed of krypton and hydrogen atoms. We have found that such samples contain very high average concentrations of hydrogen atoms (∼1018cm-3) as obtained by integration of the microwave absorption signal. Local concentrations (∼1019cm-3) of H atoms were calculated from the ESR line width. Detailed studies of the ESR line shapes lead to the conclusion that a large fraction of the H atoms lies on the krypton cluster surfaces.

Liquid Crystalline Phase Behavior of High Molecular Weight DNA: A Comparative Study of the Influence of Metal Ions of Different Size, Charge and Binding Mode

The ability of Li(+), Na(+), K(+), Rb(+), Cs(+), Mg(2+), Ca(2+), Sr(2+), Ba(2+), Cu(2+), Cd(2+), Al(3+), V(4+), Hg(2+), Pd(2+), Au(3+), and Pt(4+) to provoke liquid crystalline (LC) phases in high molecular weight DNA was investigated. The alkali and alkaline earth metal ions provoked typical cholesteric/columnar structures, whereas transition metal ions precipitated DNA into solid/translucent gel-like aggregates. Heavy metal ions reduced viscosity of DNA solution, disrupting rigid, rod-like DNA structure necessary for LC textures. Three-layer quantum mechanical-molecular mechanical (QM/MM) studies of Li(+), Na(+), K(+), Mg(2+), and Ca(2+) binding DNA fragment suggested several possible binding modes of these ions to the phosphate groups. The dianion mode of metal binding, involving the phosphate groups of both strands of DNA, allowed for higher DNA binding affinity of the alkaline earth metal ions. These results have implications in understanding the biological role of metal ions and developing DNA-based sensors and nanoelectronic devices.

SQUID Measurements of the Susceptibilities of Impurity-Helium Condensates

For the first time, the magnetic susceptibilities of impurity-helium condensates containing magnetic impurities have been measured via a SQUID magnetometer in the temperature range between 1.1 and 2.1 K. The impurity-helium condensates are gel-like aggregates of nanocrystallites composed of molecules and free radicals. Molecular oxygen nanocrystals and atomic nitrogen embedded in molecular N2 nanocrystals have been studied. The susceptibilities of the samples containing nitrogen atoms followed Curie-Weiss behavior with very small Weiss temperatures ranging from +0 to −0.4 K. The behavior of samples composed of O2 nanocrystals deviated sharply from the results for bulk solid O2. The susceptibility of the samples was 102 larger than for bulk solid O2 and showed Curie-Weiss behavior with a Weiss temperature in the range from −4.5 to −5 K. This result is qualitatively consistent with results obtained in other laboratories for O2 confined in restricted geometries.

Thermoreversible lysozyme hydrogels: properties and an insight into the gelation pathway.

The gelation behaviour of aqueous solutions of hen egg white lysozyme (HEWL) in the presence of 20 mM DTT in the concentration range 0.7 to 4.0 mM has been investigated using microDSC, FTIR, cryoTEM, SANS and oscillatory rheology. The macroscopic critical gelation concentration, Cgel, was found to be ∼ 3.0 mM. The disruption of the disulfide bonds by the DTT and the destabilisation of the protein were found to be a prerequisite for the formation of β-sheet rich fibrils under the mild conditions used in this work. Using our methodology the hydrogels obtained have a pH of 7, hence are suitable for cell culture, and are also thermoreversible. The hydrogel melting temperature was found to increase with increasing concentration and a similar structure was observed across the concentration range investigated. Our results suggest these systems are composed of a well defined regular network where single β-sheet rich fibrils (∼ 3 nm diameter) form initially, then two of these fibrils associate two-by-two to form junctions (∼ 6 nm diameter) and then on cooling further aggregate to form larger bundles of fibres. The network mesh size was found to decrease with increasing concentration. Our results suggest that below Cgel small unconnected gel-like aggregates exist that have a similar structure to the hydrogels obtained above Cgel. Using our data we propose a model for the denaturation and gelation behaviour of our system. During the first heating an α-helix to β-sheet molecular transition for the protein conformation occurs resulting in β-sheet rich fibrils forming through the self-assembly of β-sheet rich denaturated proteins. At high temperature the solution contains β-sheet rich fibrils with dissolved protein. On cooling an increase in the amount of β-sheet was observed via FTIR suggesting that as the temperature is decreased more and more protein forms β-sheet rich fibrils. At the gelation temperature these fibrils associate two-by-two to form the network junctions resulting in the macroscopic gelation of the sample. Our results suggest the network junctions are formed via specific hydrophobic interactions. The hydrogels elastic modulus was found to scale as C2.45 for C > Cgel.

Formation of Gel-Like Systems of an 2,6,8,12,14,18,20,24-Octahydroxypyridine[4]arene and an 2-Aminonaphthyridine

2,6,8,12,14,18,20,24-Octahydroxypyridine[4]arenes are capable of forming complexes with suitable partner molecules. In apolar media the formation of gel-like aggregates of an octahydroxypyridine[4]arene and an 2-aminonaphthyridine at a ratio of 1:4 was observed. This viscous liquid was investigated by means of rheology and TEM techniques and was characterized as shear thinning and thermoreversible gel-like solution which is strongly affected by changes in the stoichiometry of the complex partners. (© Wiley-VCH Verlag GmbH, 69451 Weinheim, Germany, 2002)

A novel soluble aminocellulose derivative type: its transparent film-forming properties and its efficient coupling with enzyme proteins for biosensors

By means of the reaction of 6-tosylcellulose derivatives with diamino compounds, it has been possible to provide a new soluble and film-forming aminocellulose-derivative type with a diamine residue at C-6 of the anhydroglucose unit (AGU) and with so-called solubilizing groups, such as acetate, benzoate, carbanilate, methoxy and/or tosylate groups at C-2/C-3. For example, aminocellulose derivatives were synthesized with aliphatic diamine residues of different alkyl chain lengths ((CH2)m with m = 2, 4, 6, 8, 12) at C-6. Other new aminocellulose derivatives were those with an aromatic diamine residue, e.g., 1,4-phenylenediamine or benzidine residue and others, at C-6. Depending on the structure of the diamine residue at C-6 and of the substituents at C-2/C-3, the aminocellulose derivatives were soluble in various solvents, mostly in N,N-dimethylacetamide (DMA) or dimethylsulfoxide (DMSO). Aminocelluloses with a diaminoethane or diaminobutane residue at C-6 and methoxy groups at C-2/C-3 were soluble in water. All the amino celluloses synthesized formed transparent films from their solutions. These aminocelluloses apparently form superstructures or so-called supramolecular architectures according to a structure-inherent organization principle, which became visible, for example, in the case of PDA cellulose tosylates (in DMA) as gel-like aggregates after approx. 1 week of storage at 4°C. The superstructures or aggregates could be imaged on the aminocellulose film surface by atomic force microscopy (AFM) in the form of characteristic topographic structures, e. g. as “hole” structures. In this way, structural and environment-induced factors influencing the aggregate formation were found. The transparent aminocellulose films were excellently suited for covalent coupling with oxidoreductase enzymes such as glucose oxidase (GOD), lactate oxidase (LOD), peroxidase (POD) via bifunctional compounds. A number of new bifunctional enzyme coupling reactions, e. g. via L-ascorbic acid or benzenedisulfonic dichlorides forming amide or sulfonamide coupling structures led to functionally stable nanostructure building blocks with recognition patterns in the case of GOD and POD coupling to PDA cellulose tosylate films. The PDA cellulose derivatives proved to be promising cellulose structural units because the redox-chromogenic PDA residue at C-6 provides the derivatives with a wide range of reaction possibilities, e.g. diazo coupling reactions, NH2-reactive coupling reactions and oxidative coupling reactions to redox dyes.

Chemosensitizers of the multixenobiotic resistance in amorphous aggregates (marine snow): etiology of mass killing on the benthos in the Northern Adriatic?

Periodically appearing amorphous aggregates, `marine snow', are formed in the sea and if settled as mats on the sea bottom cause death of benthic metazoans. Especially those animals are killed which are sessile filter feeders, e.g. sponges, mussels, or Anthozoa. The etiology of the toxic principle(s) is not yet well understood. Gel-like marine snow aggregates occurred in the Northern Adriatic during summer 1997. Samples of these aggregates were collected during the period July to September and the outer as well as the inner zones were analyzed for (i) cell toxicity, and (ii) chemosensitizing activity of the multixenobiotic resistance (MXR) mechanism. Organic extracts were prepared and cell toxicity was determined using mouse lymphoma cells. The experiments revealed that the major activity is seen in the center of the mats of the gel-like aggregates; a growth inhibitory activity of up to 54% (correlated to 5 ml of snow sample) was determined. The same extracts were used to determine the inhibition of the P-glycoprotein (Pgp) extrusion pump which confers the multixenobiotic resistance. The analyses were performed with cells from the sponge Suberites domuncula and with gills from the clam Corbicula fluminea in situ. Both systems have been shown to express the Pgp extrusion pump. The data show that extracts from the outer zone of the gel-like aggregate samples display pronounced inhibitory activity on the MXR extrusion pump and hence act as chemosensitizers by reversing the MXP property. These findings indicate that gel-like aggregates contain compounds in the outer zone, chemosensitizer of the Pgp extrusion pump, which lower the level of protection of metazoan animals towards dissolved compounds in their surrounding milieu, and in the center toxic compounds which are—very likely—even in the absence of chemosensitizers hazardous for the invertebrates.

Yukonite, a rare Ca-Fe arsenate, from Rçdziny (Sudetes, Poland)

Yukonite, a rare hydrated Ca-Fe arsenate, occurs at Redziny (Sudetes, Poland) in a zone of weathering of a polymetallic mineralization in dolomitic marbles. Brittle, very fine-grained, gel-like aggregates of the -mineral are dark brownish or dark brown, sometimes with a noticeable violet tint. In smaller fragments it is often translucent, purple reddish, with smooth surfaces and vitreous lustre. Yukonite originated during weathering of arsenopyrite, relics of which are sometimes present within the yukonite aggregates. It is accompanied by other arsenates (pharmacosiderite, conichalcite), arsenopyrite, quartz and talc. Yukonite has been identified by X-ray diffraction and electron-microprobe analysis.