Protein Complex
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PLoS ONE ◽  
2021 ◽  
Vol 16 (11) ◽  
pp. e0249616
Shiran Lacham-Hartman ◽  
Yulia Shmidov ◽  
Evette S. Radisky ◽  
Ronit Bitton ◽  
David B. Lukatsky ◽  

Although myriad protein–protein interactions in nature use polyvalent binding, in which multiple ligands on one entity bind to multiple receptors on another, to date an affinity advantage of polyvalent binding has been demonstrated experimentally only in cases where the target receptor molecules are clustered prior to complex formation. Here, we demonstrate cooperativity in binding affinity (i.e., avidity) for a protein complex in which an engineered dimer of the amyloid precursor protein inhibitor (APPI), possessing two fully functional inhibitory loops, interacts with mesotrypsin, a soluble monomeric protein that does not self-associate or cluster spontaneously. We found that each inhibitory loop of the purified APPI homodimer was over three-fold more potent than the corresponding loop in the monovalent APPI inhibitor. This observation is consistent with a suggested mechanism whereby the two APPI loops in the homodimer simultaneously and reversibly bind two corresponding mesotrypsin monomers to mediate mesotrypsin dimerization. We propose a simple model for such dimerization that quantitatively explains the observed cooperativity in binding affinity. Binding cooperativity in this system reveals that the valency of ligands may affect avidity in protein–protein interactions including those of targets that are not surface-anchored and do not self-associate spontaneously. In this scenario, avidity may be explained by the enhanced concentration of ligand binding sites in proximity to the monomeric target, which may favor rebinding of the multiple ligand binding sites with the receptor molecules upon dissociation of the protein complex.

eLife ◽  
2021 ◽  
Vol 10 ◽  
Dawei Wang ◽  
Zuodong Ye ◽  
Wenjie Wei ◽  
Jingting Yu ◽  
Lihong Huang ◽  

Actin filaments (F-actin) have been implicated in various steps of endosomal trafficking, and the length of F-actin is controlled by actin capping proteins, such as CapZ, which is a stable heterodimeric protein complex consisting of a and β subunits. However, the role of these capping proteins in endosomal trafficking remains elusive. Here, we found that CapZ docks to endocytic vesicles via its C-terminal actin-binding motif. CapZ knockout significantly increases the F-actin density around immature early endosomes, and this impedes fusion between these vesicles, manifested by the accumulation of small endocytic vesicles in CapZ-knockout cells. CapZ also recruits several RAB5 effectors, such as Rabaptin-5, to RAB5-positive early endosomes via its N-terminal domain, and this further activates RAB5. Collectively, our results indicate that CapZ regulates endosomal trafficking by controlling actin density around early endosomes and recruiting RAB5 effectors.

2021 ◽  
Vol 7 (1) ◽  
Shuai Lu ◽  
Yayun Gu ◽  
Yifei Wu ◽  
Shenmin Yang ◽  
Chenmeijie Li ◽  

AbstractInner dynein arm (IDA), composed of a series of protein complex, is necessary to cilia and flagella bend formation and beating. Previous studies indicated that defects of IDA protein complex result in multiple morphological abnormalities of the sperm flagellum (MMAF) and male infertility. However, the genetic causes and molecular mechanisms in the IDAs need further exploration. Here we identified two loss-of-function variants of WDR63 in both MMAF and non-obstructive azoospermia (NOA) affected cohorts. WDR63 encodes an IDA-associated protein that is dominantly expressed in testis. We next generated Wdr63-knockout (Wdr63-KO) mice through the CRISPR-Cas9 technology. Remarkably, Wdr63-KO induced decreased sperm number, abnormal flagellar morphology and male infertility. In addition, transmission electron microscopy assay showed severely disorganized “9 + 2” axoneme and absent inner dynein arms in the spermatozoa from Wdr63-KO male mice. Mechanistically, we found that WDR63 interacted with WDR78 mainly via WD40-repeat domain and is necessary for IDA assembly. Furthermore, WDR63-associated male infertility in human and mice could be overcome by intracytoplasmic sperm injection (ICSI) treatment. In conclusion, the present study demonstrates that bi-allelic variants of WDR63 cause male infertility via abnormal inner dynein arms assembly and flagella formation and can be used as a genetic diagnostic indicator for infertility males.

2021 ◽  
Weimin Lin ◽  
Xianyu Wen ◽  
Xuexin Li ◽  
Lei Chen ◽  
Wei Wei ◽  

Excessive adipogenesis caused obesity, which was a serious risk of health and led to a series of diseases, including type II diabetes (T2D) for example. Adipocyte as the basic unit of adipose tissue has emerged as one of significant target of the treatment of obesity-related metabolic syndromes by revealed its adipogenic molecular mechanism. MicroRNAs (miRNAs) have been demonstrated involving adipogenesis, and played a crucial role in the competitive endogenous RNA (ceRNA) effect. Besides that, C/EBPα as a crucial adipogenic regulator still lacked epigenetic explanation during pre-adipocyte adipogenesis. In this study, we first verified FoxO1 was one of the ceRNA of C/EBPα. They co-regulated adipogenesis through formed a protein complex that directly bound to its promoter to activate AdipoQ, and AdipoQ (Adiponectin) was a negative adipocytokines that suppressed adipogenesis, which played an important role in retaining adipogensis balance. Moreover, an adipose tissue specific enriched miRNA, miR-144 was the key regulator of the ceRNA effect between C/EBPα and FoxO1, which mediated the C/EBPα-FoxO1 complex formation, thus altered AdipoQ, furthermore regulated pre-adipocyte adipogenesis. This research will provide a new supplementary idea of the C/EBPα epigenetic role in pre-adipocyte adipogenesis.

2021 ◽  
Vol 8 ◽  
Mauro Serricchio ◽  
Peter Bütikofer

Mitochondria are essential organelles involved in cellular energy production. The inner mitochondrial membrane protein stomatin-like protein 2 (SLP-2) is a member of the SPFH (stomatin, prohibitin, flotilin, and HflK/C) superfamily and binds to the mitochondrial glycerophospholipid cardiolipin, forming cardiolipin-enriched membrane domains to promote the assembly and/or stabilization of protein complexes involved in oxidative phosphorylation. In addition, human SLP-2 anchors a mitochondrial processing complex required for proteolytic regulation of proteins involved in mitochondrial dynamics and quality control. We now show that deletion of the gene encoding the Trypanosoma brucei homolog TbSlp2 has no effect on respiratory protein complex stability and mitochondrial functions under normal culture conditions and is dispensable for growth of T. brucei parasites. In addition, we demonstrate that TbSlp2 binds to the metalloprotease TbYme1 and together they form a large mitochondrial protein complex. The two proteins negatively regulate each other’s expression levels by accelerating protein turnover. Furthermore, we show that TbYme1 plays a role in heat-stress resistance, as TbYme1 knock-out parasites displayed mitochondrial fragmentation and loss of viability when cultured at elevated temperatures. Unbiased interaction studies uncovered putative TbYme1 substrates, some of which were differentially affected by the absence of TbYme1. Our results support emerging evidence for the presence of mitochondrial quality control pathways in this ancient eukaryote.

Biomedicines ◽  
2021 ◽  
Vol 9 (11) ◽  
pp. 1649
Aleksandra A. Mamchur ◽  
Andrei V. Moiseenko ◽  
Irina S. Panina ◽  
Igor A. Yaroshevich ◽  
Sofia S. Kudryavtseva ◽  

The molecular chaperone GroEL is designed to promote protein folding and prevent aggregation. However, the interaction between GroEL and the prion protein, PrPC, could lead to pathogenic transformation of the latter to the aggregation-prone PrPSc form. Here, the molecular basis of the interactions in the GroEL–PrP complex is studied with cryo-EM and molecular dynamics approaches. The obtained cryo-EM structure shows PrP to be bound to several subunits of GroEL at the level of their apical domains. According to MD simulations, the disordered N-domain of PrP forms much more intermolecular contacts with GroEL. Upon binding to the GroEL, the N-domain of PrP begins to form short helices, while the C-domain of PrP exhibits a tendency to unfold its α2-helix. In the absence of the nucleotides in the system, these processes are manifested at the hundred nanoseconds to microsecond timescale.

mBio ◽  
2021 ◽  
Nicole Eisenhuth ◽  
Tim Vellmer ◽  
Elisa T. Rauh ◽  
Falk Butter ◽  
Christian J. Janzen

Trypanosoma brucei is a unicellular parasite that causes devastating diseases like sleeping sickness in humans and the “nagana” disease in cattle in Africa. Fundamental to the establishment and prolongation of a trypanosome infection is the parasite's ability to escape the mammalian host's immune system by antigenic variation, which relies on periodic changes of a protein surface coat.

eLife ◽  
2021 ◽  
Vol 10 ◽  
Tooba Quidwai ◽  
Jiaolong Wang ◽  
Emma A Hall ◽  
Narcis A Petriman ◽  
Weihua Leng ◽  

Intraflagellar transport (IFT) is a highly conserved mechanism for motor-driven transport of cargo within cilia, but how this cargo is selectively transported to cilia is unclear. WDR35/IFT121 is a component of the IFT-A complex best known for its role in ciliary retrograde transport. In the absence of WDR35, small mutant cilia form but fail to enrich in diverse classes of ciliary membrane proteins. In Wdr35 mouse mutants, the non-core IFT-A components are degraded and core components accumulate at the ciliary base. We reveal deep sequence homology of WDR35 and other IFT-A subunits to α and ß' COPI coatomer subunits, and demonstrate an accumulation of 'coat-less' vesicles which fail to fuse with Wdr35 mutant cilia. We determine that recombinant non-core IFT-As can bind directly to lipids and provide the first in-situ evidence of a novel coat function for WDR35, likely with other IFT-A proteins, in delivering ciliary membrane cargo necessary for cilia elongation.

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