serine residue
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Biomolecules ◽  
2022 ◽  
Vol 12 (1) ◽  
pp. 121
Author(s):  
Denise Bellotti ◽  
Adriana Miller ◽  
Magdalena Rowińska-Żyrek ◽  
Maurizio Remelli

Zrt2 is a zinc transporter of the ZIP family. It is predicted to be located in the plasma membrane and it is essential for Candida albicans zinc uptake and growth at acidic pH. Zrt2 from C. albicans is composed of 370 amino acids and contains eight putative transmembrane domains and an extra-membrane disordered loop, corresponding to the amino acid sequence 126–215. This protein region contains at least three possible metal binding motifs: HxHxHxxD (144–153), HxxHxxEHxD (181–193) and the Glu- and Asp- rich sequence DDEEEDxE (161–168). The corresponding model peptides, protected at their termini (Ac-GPHTHSHFGD-NH2, Ac-DDEEEDLE-NH2 and Ac-PSHFAHAQEHQDP-NH2), have been investigated in order to elucidate the thermodynamic and coordination properties of their Zn2+ and Cu2+ complexes, with the further aim to identify the most effective metal binding site among the three fragments. Furthermore, we extended the investigation to the peptides Ac-GPHTHAHFGD-NH2 and Ac-PAHFAHAQEHQDP-NH2, where serine residues have been substituted by alanines in order to check if the presence of a serine residue may favor the displacement of amidic protons by Cu2+. In the native Zrt2 protein, the Ac-GPHTHSHFGD-NH2 region of the Zrt2 loop has the highest metal binding affinity, showing that three alternated histidines separated by only one residue (-HxHxH-) bind Zn2+ and Cu2+ more strongly than the region in which three histidines are separated by two and three His residues (-HxxHxxxH- in Ac-PSHFAHAQEHQDP-NH2). All studied Zrt2 loop fragments have lower affinity towards Zn2+ than the zinc(II) binding site on the Zrt1 transporter; also, all three Zrt2 regions bind Zn2+ and Cu2+ with comparable affinity below pH 5 and, therefore, may equally contribute to the metal acquisition under the most acidic conditions in which the Zrt2 transporter is expressed.


2021 ◽  
Author(s):  
Devanand T ◽  
Susmita Ghosh ◽  
Prasanna Venkatraman ◽  
Satyavani Vemparala

The effect of phosphorylation of a serine residue in the Rap protein, residing at the complex interface of Rap-Raf complex is studied using atomistic molecular dynamics simulations. As the phosphosite of interest (SER39) is buried at the interface of the Rap-Raf complex, phosphorylation of only Rap protein was simulated and then complexed with the RBD of Raf for further analysis of complex stability. Our simulations reveal that the phosophorylation increases the binding of complex through strong electrostatic interactions and changes the charge distribution of the interface significantly. This is manifested as an increase in stable salt-bridge interactions between the Rap and Raf of the complex. Network analysis clearly shows that the phosphorylation of SER39 reorganizes the community network to include the entire region of Raf chain, including, Raf L4 loop potentially affecting downstream signalling.


2021 ◽  
Author(s):  
Chen Zhu ◽  
Yi Wang ◽  
Zhen Lin ◽  
Rongxia Li ◽  
Bei Jia ◽  
...  

Abstract Clade A PP2C phosphatases are central components of the ABA-receptor coupled core signaling pathway, and are involved in multiple stress responses and developmental processes. However, the direct targets or partner proteins of A clade PP2Cs that participate in these biological processes are largely unknown. Here, we used a TurboID-based proximity labeling method to identify putative associated proteins of one A clade PP2C phosphatase, ABI1. By combining the results from affinity purification or proximity labeling of biotinylated proteins, we identified more than four hundred putative ABI1-associated proteins, including dozens of known ABI1-intreacting proteins, as well as proteins involved in TOR signaling, phospho-regulation, and other biological processes. We found that RAFs, a group of protein kinases that phosphorylate and activate SnRK2s in ABA and osmotic stress signaling, are direct substrates of ABI1. A conserved serine residue located in the P-loop of the kinase domain, corresponding to Ser619 in RAF3, is a major functional ABI1 target site. ABI1-mediated dephosphorylation on this site strongly promotes the kinase activity of most B2 and B3 RAFs. Thus, ABI1 has dual functions in ABA signaling by dephosphorylating and inhibiting SnRK2 to prevent SnRK2 activation in unstressed conditions, while dephosphorylating B2 and B3 subgroup RAFs to maintain their basal kinase activity. PP2C-mediated dephosphorylation at the conserved serine residue may be a mechanism for RAF activation in both plants and animals, with potential implications for tumorigenesis in humans.


2021 ◽  
Vol 12 ◽  
Author(s):  
Jianwei Zhou ◽  
Yonghui Qiu ◽  
Ning Zhu ◽  
Linyi Zhou ◽  
Beining Dai ◽  
...  

Porcine circovirus type 4 (PCV4) is an emerging etiological agent which was first detected in 2019. The nucleolar localization signal (NoLS) of PCV4 Cap protein and its binding host cellular proteins are still not elucidated. In the present study, we discovered a distinct novel NoLS of PCV4 Cap, which bound to the nucleolar phosphoprotein nucleophosmin-1 (NPM1). The NoLS of PCV4 Cap and serine-48 residue at the N-terminal oligomerization domain of NPM1 were necessary for PCV4 Cap/NPM1 interaction. Furthermore, the charge property of serine residue at position 48 of the NPM1 was crucial for its oligomerization and interaction with PCV4 Cap. In summary, our findings show for the first time that the PCV4 Cap NoLS and the NPM1 oligomerization determine the interaction of Cap/NPM1.


2021 ◽  
Author(s):  
Logan P. Poole ◽  
Althea Bock-Hughes ◽  
Damian E. Berardi ◽  
Kay F. Macleod

Abstract UNC51-like kinase-1 (ULK1) is the catalytic component of the autophagy pre-initiation complex that stimulates autophagy via phosphorylation of ATG14, BECLN1 and other autophagy proteins. ULK1 has also been shown to specifically promote mitophagy but the mechanistic basis of how has remained unclear. Here we show that ULK1 phosphorylates the BNIP3 mitochondrial cargo receptor on a critical serine residue (S17) adjacent to its amino terminal LIR motif. ULK1 similarly phosphorylates BNIP3L on S35. Phosphorylation of BNIP3 on S17 by ULK1 promotes interaction with LC3 and mitophagy. ULK1 interaction also promotes BNIP3 protein stability by limiting its turnover at the proteasome. The ability of ULK1 to regulate BNIP3 protein stability depends on an intact “BH3” domain and deletion of its “BH3” domain reduces BNIP3 turnover and increases BNIP3 protein levels independent of ULK1. In summary ULK1 promotes mitophagy by both stabilization of BNIP3 protein and via phosphorylation of S17 to stimulate interaction with LC3.


2021 ◽  
Vol 8 ◽  
Author(s):  
Jackson T. Baumgartner ◽  
Thahani S. Habeeb Mohammad ◽  
Mateusz P. Czub ◽  
Karolina A. Majorek ◽  
Xhulio Arolli ◽  
...  

Enzymes in the Gcn5-related N-acetyltransferase (GNAT) superfamily are widespread and critically involved in multiple cellular processes ranging from antibiotic resistance to histone modification. While acetyl transfer is the most widely catalyzed reaction, recent studies have revealed that these enzymes are also capable of performing succinylation, condensation, decarboxylation, and methylcarbamoylation reactions. The canonical chemical mechanism attributed to GNATs is a general acid/base mechanism; however, mounting evidence has cast doubt on the applicability of this mechanism to all GNATs. This study shows that the Pseudomonas aeruginosa PA3944 enzyme uses a nucleophilic serine residue and a hybrid ping-pong mechanism for catalysis instead of a general acid/base mechanism. To simplify this enzyme’s kinetic characterization, we synthesized a polymyxin B substrate analog and performed molecular docking experiments. We performed site-directed mutagenesis of key active site residues (S148 and E102) and determined the structure of the E102A mutant. We found that the serine residue is essential for catalysis toward the synthetic substrate analog and polymyxin B, but the glutamate residue is more likely important for substrate recognition or stabilization. Our results challenge the current paradigm of GNAT mechanisms and show that this common enzyme scaffold utilizes different active site residues to accomplish a diversity of catalytic reactions.


2021 ◽  
Vol 22 ◽  
Author(s):  
Arindam Maity ◽  
Debanjan Sen ◽  
Chandi Charan Kandar

: Glycogen synthase kinase -3 (GSK-3) is a protein kinase containing threonine or serine amino acid residues. GSK-3 was first discovered in 1980 as a regulatory protein kinase for its namesake, Glycogen synthase (GS) enzyme, which is responsible for the conversion of glycogen from glucose with the help of uridine diphosphate glucose (UDP-Glu) residue. GSK-3 has two isoforms present in human’s beings namely GSK-3 α (serine residue at 21 position) and GSK-3 β (serine residue at 9 position). GSK-3 has two terminals namely C- terminal and N- terminal. C-terminal of GSK-3 looks like α- helix conformation which acts as activator loop and is responsible for helping to position residues like symphoric effect involved in ATP binding and catalysis of substrates. On the other hand, the N- terminal of GSK-3 is like β- strand conformation which acts as an inhibitory loop, and having tyrosine molecule at 216 positions essential for showing the complete GSK-3 activity. N- terminal of GSK-3 is responsible for ATP binding activity and exhibits various biological activities like cell signaling, gene induction following activation of T cell receptor, apoptosis, protein translation, glycogen metabolism, and inflammatory process. Activation of GSK-3 causes Pro- inflammatory actions, increase the binding activity of NF-kB (pro-inflammatory genes), increase the transactivation activity of NF-kB, Increase the phosphorylation of p105, decrease the transactivation activity of C/EBPβ (anti-inflammatory genes) resulting in a large number of prevalent diseases such as diabetes, cancer, neurodegenerative diseases, psychiatric diseases, mood disorders, etc. Glycogen synthase kinase inhibitors (GSK-3 inhibitors) are various chemotypes and have a different mechanism of actions. They are obtained from different sources such as natural products, synthetic ATP as well as non-ATP competitive inhibitors along with substratecompetitive inhibitors. The inhibitors of GSK3 proved to be providing very potent anti-inflammatory action. GSK-3 inhibitors are useful in treating different prevalent disorders, such as neurodegenerative diseases including Alzheimer's disease, hyperglycemia, cancer disease, and mood disorders like depression, etc. In this review, we have highlighted the views regarding the description and types of GSK, inflammation process, and the factors affecting inflammation, the relationship between inflammation and GSK, GSK3 inhibitors, and finally, the reflection of various natural as well as synthetic GSK3 inhibitors having anti-inflammatory activity.


2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Ai-Ling Tian ◽  
Qi Wu ◽  
Peng Liu ◽  
Liwei Zhao ◽  
Isabelle Martins ◽  
...  

AbstractThe integrated stress response manifests with the phosphorylation of eukaryotic initiation factor 2α (eIF2α) on serine residue 51 and plays a major role in the adaptation of cells to endoplasmic reticulum stress in the initiation of autophagy and in the ignition of immune responses. Here, we report that lysosomotropic agents, including azithromycin, chloroquine, and hydroxychloroquine, can trigger eIF2α phosphorylation in vitro (in cultured human cells) and, as validated for hydroxychloroquine, in vivo (in mice). Cells bearing a non-phosphorylatable eIF2α mutant (S51A) failed to accumulate autophagic puncta in response to azithromycin, chloroquine, and hydroxychloroquine. Conversely, two inhibitors of eIF2α dephosphorylation, nelfinavir and salubrinal, enhanced the induction of such autophagic puncta. Altogether, these results point to the unexpected capacity of azithromycin, chloroquine, and hydroxychloroquine to elicit the integrated stress response.


2020 ◽  
Vol 21 (23) ◽  
pp. 8971
Author(s):  
Martin S. King ◽  
Sotiria Tavoulari ◽  
Vasiliki Mavridou ◽  
Alannah C. King ◽  
John Mifsud ◽  
...  

Cryptosporidiumparvum is a clinically important eukaryotic parasite that causes the disease cryptosporidiosis, which manifests with gastroenteritis-like symptoms. The protist has mitosomes, which are organelles of mitochondrial origin that have only been partially characterized. The genome encodes a highly reduced set of transport proteins of the SLC25 mitochondrial carrier family of unknown function. Here, we have studied the transport properties of one member of the C. parvum carrier family, demonstrating that it resembles the mitochondrial ADP/ATP carrier of eukaryotes. However, this carrier has a broader substrate specificity for nucleotides, transporting adenosine, thymidine, and uridine di- and triphosphates in contrast to its mitochondrial orthologues, which have a strict substrate specificity for ADP and ATP. Inspection of the putative translocation pathway highlights a cysteine residue, which is a serine in mitochondrial ADP/ATP carriers. When the serine residue is replaced by cysteine or larger hydrophobic residues in the yeast mitochondrial ADP/ATP carrier, the substrate specificity becomes broad, showing that this residue is important for nucleotide base selectivity in ADP/ATP carriers.


2020 ◽  
Vol 27 ◽  
Author(s):  
Guo-Ying Qian ◽  
Gyutae Lim ◽  
Shang-Jun Yin ◽  
Jun-Mo Yang ◽  
Jinhyuk Lee ◽  
...  

Background: Background: Fibrinolytic protease from Euphausia superba (EFP) was isolated. Objective: Biochemical distinctions, regulation of the catalytic function, and the key residues of EFP were investigated. Methods: The serial inhibition kinetic evaluations coupled with measurements of fluorescence spectra in the presence of 4- (2-aminoethyl) benzene sulfonyl fluoride hydrochloride (AEBSF) was conducted. The computational molecular dynamics (MD) simulations were also applied for a comparative study. Results: The enzyme behaved as a monomeric protein with a molecular mass of about 28.6 kD with Km BApNA = 0.629 ± 0.02 mM and kcat/Km BApNA = 7.08 s-1 /mM. The real-time interval measurements revealed that the inactivation was a first-order reaction, with the kinetic processes shifting from a monophase to a biphase. Measurements of fluorescence spectra showed that serine residue modification by AEBSF directly caused conspicuous changes of the tertiary structures and exposed hydrophobic surfaces. Some osmolytes were applied to find protective roles. These results confirmed that the active region of EFP is more flexible than the overall enzyme molecule and serine, as the key residue, is associated with the regional unfolding of EFP in addition to its catalytic role. The MD simulations were supportive to the kinetics data. Conclusion: Our study indicated that EFP has an essential serine residue for its catalyst function and associated folding behaviors. Also, the functional role of osmolytes such as proline and glycine that may play a role in defense mechanisms from environmental adaptation in a krill’s body was suggested.


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