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Four new polyacetylene amides, siphonellamides A-D (1-4), and one new fatty amide, siphonellamide E (5), together with a known indole fatty amide (6) and callyspongamide A (7), were isolated from the Red Sea marine sponge Siphonochalina siphonella. The structures of 1-5 were elucidated by extensive analyses of their 1D- and 2D-NMR spectra and MS. The isolated compounds were assessed for their cytotoxicity against HeLa, MCF-7, and A549 cancer cell lines. Compounds 1 and 2 exhibited cytotoxic activities with IC 50 values ranging from 9.4 to 34.1 μM, while 5 was only cytotoxic to HeLa cells, with an IC 50 value of 78.4 μM. Compound 7 showed moderate cytotoxicity against all tested cell lines., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflicts of interest., (Copyright © 2020 Elsevier B.V. All rights reserved.)

The highly sulfated domains of heparan sulfate (HS), alias HS S-domains, are made up of repeated trisulfated disaccharide units [iduronic acid (2S)-glucosamine (NS, 6S)] and are selectively remodeled by extracellular endoglucosamine 6-sulfatases (Sulfs). Although HS S-domains are critical for signal transduction of several growth factors, their roles in amyloidoses are not yet fully understood. Herein, we found HS S-domains in the kidney of a patient with transthyretin amyloidosis. In in vitro assays with cells stably expressing human Sulfs, heparin, a structural analog of HS S-domains, promoted aggregation of transthyretin in an HS S-domain-dependent manner. Interactions of cells with transthyretin fibrils and cytotoxicity of these fibrils also depended on HS S-domains at the cell surface. Furthermore, glypican-5, encoded by the susceptibility gene for nephrotic syndrome GPC5, was found to be accumulated in the transthyretin amyloidosis kidney. Our study, thus, provides a novel insight into the pathologic roles of HS S-domains in amyloidoses, and we propose that enzymatic remodeling of HS chains by Sulfs may offer an effective approach to inhibiting formation and cytotoxicity of amyloid fibrils., (Copyright © 2019 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

In eukaryotes, RNA polymerase II requires general transcription factors to initiate mRNA transcription. TFIIE subunits α and β form a heterodimer and recruit TFIIH to complete the assembly of the pre-initiation complex. Here, we have determined the crystal structure of human TFIIE at atomic resolution. The N-terminal half of TFIIEα forms an extended winged helix (WH) domain with an additional helix, followed by a zinc-finger domain. TFIIEβ contains the WH2 domain, followed by two coiled-coil helices intertwining with TFIIEα. We also showed that TFIIEα binds to TFIIEβ with nanomolar affinity using isothermal titration calorimetry. In addition, mutations on the residues involved in the interactions resulted in severe growth defects in yeast. Lack of the C-terminal region of yeast TFIIEβ causes a mild growth defect in vivo. These findings provide a structural basis for understanding the functional mechanisms of TFIIE in the context of pre-initiation complex formation and transcription initiation., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

The endosomal sorting complex required for transport (ESCRT) facilitates roles in membrane remodeling, such as multivesicular body biogenesis, enveloped virus budding and cell division. In yeast, Vps4 plays a crucial role in intraluminal vesicle formation by disassembling ESCRT proteins. Vps4 is recruited by ESCRT-III proteins to the endosomal membrane through the interaction between the microtubule interacting and trafficking (MIT) domain of Vps4 and the C-terminal MIT-interacting motif (MIM) of ESCRT-III proteins. Here, we have determined the crystal structure of Vps4-MIT in a complex with Vps20, a member of ESCRT-III, and revealed that Vps20 adopts a unique MIM2 conformation. Based on structural comparisons with other known MIM2s, we have refined the consensus sequence of MIM2. We have shown that another ESCRT-III protein, Ist1, binds to Vps4-MIT via its C-terminal MIM1 with higher affinity than Vps2, but lacks MIM2 by surface plasmon resonance. Surprisingly, the Ist1 MIM1 competed with the MIM2 of Vfa1, a regulator of Vps4, for binding to Vps4-MIT, even though these MIMs bind in non-overlapping sites on the MIT. These findings provide insight into the allosteric recognition of MIMs of ESCRT-III by Vps4 and also the regulation of ESCRT machinery at the last step of membrane remodeling., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

D-serine is an endogenous coagonist of the N-methyl-D-aspartate (NMDA)-type glutamate receptor in the central nervous system and its synthesis is catalyzed by serine racemase (SR). Recently, the NMDA receptor has been found to be expressed in keratinocytes (KCs) of the skin and involved in the regulation of KC growth and differentiation. However, the localization and role of SR in the skin remain unknown. Here, using SR-knockout (SR-KO) mice as the control, we demonstrated the localization of the SR protein in the granular and cornified layer of the epidermis of wild-type (WT) mice and its appearance in confluent WT KCs. We also demonstrated the existence of a mechanism for conversion of L-serine to D-serine in epidermal KCs. Furthermore, we found increased expression levels of genes involved in the differentiation of epidermal KCs in adult SR-KO mice, and alterations in the barrier function and ultrastructure of the epidermis in postnatal day 5 SR-KO mice. Our findings suggest that SR in the skin epidermis is involved in the differentiation of epidermal KCs and the formation of the skin barrier.

Aims: Transthyretin (TTR)-related familial amyloidotic polyneuropathy (FAP) is characterized by the systemic accumulation of amyloid fibrils caused by amyloidogenic. Our previous studies demonstrated that albumin played a role in the inhibition of TTR amyloid-formation. The aim of this study was to evaluate the effect of albumin on TTR disposition and tissue deposition in vivo., Main Methods: For pharmacokinetic studies, recombinant wild-type TTR (rTTR) and recombinant amyloidogenic TTR Val30Met (rATTR V30M) were labeled with iodine and administered to Sprague-Dawley rats and analbuminemia rats (NAR: Nagase Analbuminemia Rats). The deposition of ATTR V30M was also analyzed by immunohistochemistry in the transgenic (Tg) rats possessing a human ATTR V30M gene (ATTR V30M Tg rats) and NAR possessing a human ATTR V30M gene (ATTR V30M Tg NAR)., Key Findings: The presence of albumin had no effect on the tissue distribution of either rTTR or rATTR V30M. However, more ATTR V30M was deposited in the hearts, stomachs and small intestines of ATTR V30M Tg NAR rats, compared to ATTR V30M Tg rats., Significance: Although the disposition of TTR and ATTR V30M was unaffected by the presence of albumin, the deposition of ATTR V30M in some organs was apparently increased in the absence of albumin compared to the presence of albumin. These results show that albumin would contribute to suppressing the tissue deposition of TTR in pathogenesis of FAP, but does not affect the disposition of TTR., (Copyright © 2013 Elsevier Inc. All rights reserved.)

HAMLET (Human Alpha-lactalbumin Made LEthal to Tumor cells), which was identified in human breast milk as an alpha-lactalbumin (LA)-oleic acid complex, kills tumor cells, selectively. Although it may have potential as a therapeutic agent against various tumor cells, only low-volume methods for its production exist. In this study, heat treatment was used to produce complexes from LAs and oleic acid using a simple method. In the case of human LA and oleic acid, heat-treated samples apparently showed much stronger activities than those treated at room temperature, with cytotoxicities equal to that of HAMLET. Furthermore, circular dichroism spectroscopy revealed that heat-treated samples lost their tertiary structure, suggesting a molten globule as oleic acid-bound LA. BLA samples also showed strong activities by heat treatment. Batch production with heat treatment can efficiently convert LAs into tumoricidal complexes.

Bacteriophage Mu is a double-stranded DNA phage that consists of an icosahedral head, a contractile tail with baseplate and six tail fibers, similar to the well-studied T-even phages. The baseplate of bacteriophage Mu, which recognizes and attaches to a host cell during infection, consists of at least eight different proteins. The baseplate protein, gp44, is essential for bacteriophage Mu assembly and the generation of viable phages. To investigate the role of gp44 in baseplate assembly and infection, the crystal structure of gp44 was determined at 2.1A resolution by the multiple isomorphous replacement method. The overall structure of the gp44 trimer is similar to that of the T4 phage gp27 trimer, which forms the central hub of the T4 baseplate, although these proteins share very little primary sequence homology. Based on these data, we confirm that gp44 exists as a trimer exhibiting a hub-like structure with an inner diameter of 25A through which DNA can presumably pass during infection. The molecular surface of the gp44 trimer that abuts the host cell membrane is positively charged, and it is likely that Mu phage interacts with the membrane through electrostatic interactions mediated by gp44.

At low ionic strength, apoplastocyanin forms an unfolded state under non-denaturing conditions. The refolding of this state is sufficiently slow to allow real-time NMR experiments to be performed. Folding of apoplastocyanin, initiated by the addition of salt and followed by real-time 2D 1H-15N heteronuclear single quantum coherence (HSQC) spectroscopy, is highly cooperative. A concomitant increase in the intensity of both sequential and long-range nuclear Overhauser effects (NOEs) between backbone amide protons in successive acquisitions of 1H-15N HSQC-NOESY-HSQC spectra provides the first direct observation of the development of structure-specific NOEs as a protein folds. Our results show that the local and long-range interactions in the native apoplastocyanin are formed simultaneously, consistent with highly cooperative formation of the native structure.