Biochemistry Biochemistry最新文献_第9页

Biochemical, Structural, and Conformational Characterization of a Fungal Ethylene-Forming Enzyme.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-03-07 DOI: 10.1021/acs.biochem.5c00038

Shramana Chatterjee, Joel A Rankin, Mark A Farrugia, Simahudeen Bathir J S Rifayee, Christo Z Christov, Jian Hu, Robert P Hausinger

{"title":"Biochemical, Structural, and Conformational Characterization of a Fungal Ethylene-Forming Enzyme.","authors":"Shramana Chatterjee, Joel A Rankin, Mark A Farrugia, Simahudeen Bathir J S Rifayee, Christo Z Christov, Jian Hu, Robert P Hausinger","doi":"10.1021/acs.biochem.5c00038","DOIUrl":"https://doi.org/10.1021/acs.biochem.5c00038","url":null,"abstract":"The ethylene-forming enzyme (EFE) from the fungus Penicillium digitatum strain Pd1 was heterologously produced in Escherichia coli and its properties were compared to the extensively characterized bacterial enzyme from Pseudomonas savastanoi strain PK2. Both enzymes catalyze four reactions: the conversion of 2-oxoglutarate (2OG) to ethylene and CO2, oxidative decarboxylation of 2OG coupled to l-arginine (l-Arg) hydroxylation, uncoupled oxidative decarboxylation of 2OG, and the production of 3-hydroxypropionate (3-HP) from 2OG. The strain Pd1 enzyme exhibited a greater ratio of ethylene production over l-Arg hydroxylation than the PK2 strain EFE. The uncoupled decarboxylation of 2OG and 3-HP production are minor reactions in both cases, but they occur to a greater extent using the fungal enzyme. Additional distinctions of the fungal versus bacterial enzyme are noted in the absorbance maxima and l-Arg dependence of their anaerobic electronic spectra. The structures of the Pd1 EFE apoprotein and the EFE·Mn(II)·2OG complex resembled the corresponding structures of the PK2 enzyme, but notable structural differences were observed in the computationally predicted Pd1 EFE·Fe(II)·2OG·l-Arg complex versus the PK2 EFE·Mn(II)·2OG·l-Arg crystal structure. These studies extend our biochemical understanding and represent the first structural and conformational characterization of a eukaryotic EFE.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":""},"PeriodicalIF":2.9,"publicationDate":"2025-03-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143571550","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Crystal Structure, Modeling, and Identification of Key Residues Provide Insights into the Mechanism of the Key Toxoflavin Biosynthesis Protein ToxD

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-03-06 DOI: 10.1021/acs.biochem.4c0042110.1021/acs.biochem.4c00421

Savannah F. Justen, Michael K. Fenwick, Kyle K. Axt, James A. Cherry, Steven E. Ealick and Benjamin Philmus*,

{"title":"Crystal Structure, Modeling, and Identification of Key Residues Provide Insights into the Mechanism of the Key Toxoflavin Biosynthesis Protein ToxD","authors":"Savannah F. Justen, Michael K. Fenwick, Kyle K. Axt, James A. Cherry, Steven E. Ealick and Benjamin Philmus*, ","doi":"10.1021/acs.biochem.4c0042110.1021/acs.biochem.4c00421","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00421https://doi.org/10.1021/acs.biochem.4c00421","url":null,"abstract":"Toxoflavin, a toxic secondary metabolite produced by a variety of bacteria, has been implicated as a causative agent in food poisoning and a virulence factor in phytopathogenic bacteria. This toxin is produced by genes encoded in the tox operon in Burkholderia glumae, in which the encoded protein, ToxD, was previously characterized as essential for toxoflavin production. To better understand the function of ToxD in toxoflavin biosynthesis and provide a basis for future work to develop inhibitors of ToxD, we undertook the identification of structurally and catalytically important amino acid residues through a combination of X-ray crystallography and site directed mutagenesis. We solved the structure of BgToxD, which crystallized as a dimer, to 1.8 Å resolution. We identified a citrate molecule in the putative active site. To investigate the role of individual residues, we used Pseudomonas protegens Pf-5, a BL1 plant protective bacterium known to produce toxoflavin, and created mutants in the ToxD-homologue PFL1035. Using a multiple sequence alignment and the BgToxD structure, we identified and explored the functional importance of 12 conserved residues in the putative active site. Eight variants of PFL1035 resulted in no observable production of toxoflavin. In contrast, four ToxD variants resulted in reduced but detectable toxoflavin production suggesting a nonessential role. The crystal structure and structural models of the substrate and intermediate bound enzyme provide a molecular interpretation for the mutagenesis data.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":"64 6","pages":"1199–1211 1199–1211"},"PeriodicalIF":2.9,"publicationDate":"2025-03-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143635886","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Alanine Scanning to Define Membrane Protein–Lipid Interaction Sites Using Native Mass Spectrometry

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-03-06 DOI: 10.1021/acs.biochem.4c0071710.1021/acs.biochem.4c00717

Hiruni S. Jayasekera, Farhana Afrin Mohona, Madison J. De Jesus, Katherine M. Miller and Michael T. Marty*,

{"title":"Alanine Scanning to Define Membrane Protein–Lipid Interaction Sites Using Native Mass Spectrometry","authors":"Hiruni S. Jayasekera, Farhana Afrin Mohona, Madison J. De Jesus, Katherine M. Miller and Michael T. Marty*, ","doi":"10.1021/acs.biochem.4c0071710.1021/acs.biochem.4c00717","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00717https://doi.org/10.1021/acs.biochem.4c00717","url":null,"abstract":"Lipids surrounding membrane proteins interact with different sites on the protein with varying specificities, ranging from highly specific to weak interactions. These interactions can modulate the structure, function, and stability of membrane proteins. Thus, to better understand membrane protein structure and function, it is important to identify the locations of lipid binding and the relative specificities of lipid binding at these sites. In our previous native mass spectrometry (MS) study, we developed a single and double mutant analysis approach to profile the contribution of specific residues toward lipid binding. Here, we extend this method by screening a broad range of mutants of AqpZ to identify specific lipid binding sites and by measuring binding of different lipid types to measure the selectivity of different lipids at selected binding sites. We complemented these native MS studies with molecular dynamics (MD) simulations to visualize lipid interactions at selected sites. We discovered that AqpZ is selective toward cardiolipins (CL) but only at specific sites. Specifically, CL orients with its headgroup facing the cytoplasmic side, and its acyl chains interact with a hydrophobic pocket located at the monomeric interface within the lipid bilayer. Overall, this integrative approach provides unique insights into lipid binding sites and the selectivity of various lipids toward AqpZ, enabling us to map the AqpZ protein structure based on the lipid affinity.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":"64 6","pages":"1308–1316 1308–1316"},"PeriodicalIF":2.9,"publicationDate":"2025-03-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143636013","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Unveiling the Catalytic Mechanism of Abl1 Kinase: A Single-Magnesium Ion Pathway for Phosphoryl Transfer 揭示 Abl1 激酶的催化机制：磷酰转移的单镁离子途径

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-03-05 DOI: 10.1021/acs.biochem.4c0083810.1021/acs.biochem.4c00838

Sinisa Bjelic*, Stella Hernandez Maganhi and Ran Friedman,

{"title":"Unveiling the Catalytic Mechanism of Abl1 Kinase: A Single-Magnesium Ion Pathway for Phosphoryl Transfer","authors":"Sinisa Bjelic*, Stella Hernandez Maganhi and Ran Friedman, ","doi":"10.1021/acs.biochem.4c0083810.1021/acs.biochem.4c00838","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00838https://doi.org/10.1021/acs.biochem.4c00838","url":null,"abstract":"Abl1, a nonreceptor tyrosine kinase closely related to Src kinase, regulates critical cellular processes like proliferation, differentiation, cytoskeletal dynamics, and response to environmental cues through phosphorylation-driven activation. Dysregulation places it centrally in the oncogenic pathway leading to blood cancers. making it an ideal drug target for small molecule inhibitors. We sought to understand the underlying mechanism of the phosphoryl-transfer step from the ATP molecule to the substrate tyrosine, as carried out by the Abl1 enzyme. By calculating free energy profiles for the reaction using the empirical valence bond representation of the reacting fragments paired with molecular dynamics and free energy perturbation calculations, a combination of several plausible reaction pathways, ATP conformations, and the number of magnesium ion cofactors have been investigated. For the best-catalyzed pathway, which proceeds through a dissociative mechanism with a single magnesium ion situated in Site I, a close agreement was reached with the experimentally determined catalytic rates. We conclude that the catalytic mechanism in Abl1 requires one magnesium ion for efficient catalysis, unlike other kinases, where two ions are utilized. A better overall understanding of the phosphoryl-transfer reactions in Abl1 can be used for type-I inhibitor development and generally contributes to a comprehensive overview of the mechanism for ATP-driven reactions.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":"64 6","pages":"1415–1424 1415–1424"},"PeriodicalIF":2.9,"publicationDate":"2025-03-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.acs.org/doi/epdf/10.1021/acs.biochem.4c00838","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143635953","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Controlled Enzyme Cargo Loading in Engineered Bacterial Microcompartment Shells

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-03-05 DOI: 10.1021/acs.biochem.4c0070910.1021/acs.biochem.4c00709

Nicholas M. Tefft, Yali Wang, Alexander Jussupow, Michael Feig and Michaela A. TerAvest*,

{"title":"Controlled Enzyme Cargo Loading in Engineered Bacterial Microcompartment Shells","authors":"Nicholas M. Tefft, Yali Wang, Alexander Jussupow, Michael Feig and Michaela A. TerAvest*, ","doi":"10.1021/acs.biochem.4c0070910.1021/acs.biochem.4c00709","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00709https://doi.org/10.1021/acs.biochem.4c00709","url":null,"abstract":"Bacterial microcompartments (BMCs) are nanometer-scale organelles with a protein-based shell that serve to colocalize and encapsulate metabolic enzymes. They may provide a range of benefits to improve pathway catalysis, including substrate channeling and selective permeability. Several groups are working toward using BMC shells as a platform for enhancing engineered metabolic pathways. The microcompartment shell of Haliangium ochraceum (HO) has emerged as a versatile and modular shell system that can be expressed and assembled outside its native host and with non-native cargo. Further, the HO shell has been modified to use the engineered protein conjugation system SpyCatcher–SpyTag for non-native cargo loading. Here, we used a model enzyme, triose phosphate isomerase (Tpi), to study non-native cargo loading into four HO shell variants and begin to understand maximal shell loading levels. We also measured activity of Tpi encapsulated in the HO shell variants and found that activity was determined by the amount of cargo loaded and was not strongly impacted by the predicted permeability of the shell variant to large molecules. All shell variants tested could be used to generate active, Tpi-loaded versions, but the simplest variants assembled most robustly. We propose that the simple variant is the most promising for continued development as a metabolic engineering platform.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":"64 6","pages":"1285–1292 1285–1292"},"PeriodicalIF":2.9,"publicationDate":"2025-03-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.acs.org/doi/epdf/10.1021/acs.biochem.4c00709","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143635961","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Multiple Copper Ions Bind to and Promote the Oligomerization of Huntingtin Protein with Nonpathological Repeat Expansions.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-03-04 Epub Date: 2025-02-21 DOI: 10.1021/acs.biochem.5c00012

Deepa Neupane, Miguel Santos-Fernandez, Francisco Fernandez-Lima, Katlyn K Meier

{"title":"Multiple Copper Ions Bind to and Promote the Oligomerization of Huntingtin Protein with Nonpathological Repeat Expansions.","authors":"Deepa Neupane, Miguel Santos-Fernandez, Francisco Fernandez-Lima, Katlyn K Meier","doi":"10.1021/acs.biochem.5c00012","DOIUrl":"10.1021/acs.biochem.5c00012","url":null,"abstract":"Huntington's disease (HD) is a fatal neurodegenerative disease characterized by the expression of huntingtin protein (htt) that has a polyglutamine (CAG; polyQ) repeat domain consisting of 36 or more glutamines (mhtt). Historically, mhtt is more broadly associated with HD severity, as are elevated metal levels observed in HD patients. The depletion of wild-type (WT) htt (fewer than 36Qs) is also recognized as a contributing factor to HD progression; however, many questions remain about the interactions of biorelevant metals with WT htt and the impact of the interactions on protein aggregation. In the present work, we utilize a combination of biochemical assays and spectroscopic techniques to provide insights into the interaction of copper with an in vitro htt model (N171-17Q). Herein, we use sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and dynamic light scattering to show that the addition of equimolar or higher concentrations of Cu(II) to htt induces time- and temperature-dependent protein oligomerization/aggregation. Additionally, chelation assays, trapped ion mobility spectrometry, and mass spectrometry confirm the (i) rapid reduction of Cu(II) in the presence of N171-17Q htt, (ii) direct binding of multiple copper ions per protein, and (iii) complex Cu:htt speciation profile with a preference for three distinct Cu:htt states. These findings contribute to our molecular level understanding of copper's role in the depletion and oligomerization/aggregation of WT htt while underscoring the physiological significance of our work, its potential relevance to metal binding in mhtt, and its significance for identifying new avenues for biomarker exploration and therapeutic design strategies.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":"1121-1135"},"PeriodicalIF":2.9,"publicationDate":"2025-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143472008","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Aβ40 Fibril Assembly on Human Cerebral Smooth Muscle Cells Impairs Cell Viability.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-03-04 Epub Date: 2025-01-06 DOI: 10.1021/acs.biochem.4c00478

Brandon Irizarry, Judianne Davis, Jitika Rajpoot, Xiaoyue Zhu, Feng Xu, Steven O Smith, William E Van Nostrand

{"title":"Aβ40 Fibril Assembly on Human Cerebral Smooth Muscle Cells Impairs Cell Viability.","authors":"Brandon Irizarry, Judianne Davis, Jitika Rajpoot, Xiaoyue Zhu, Feng Xu, Steven O Smith, William E Van Nostrand","doi":"10.1021/acs.biochem.4c00478","DOIUrl":"10.1021/acs.biochem.4c00478","url":null,"abstract":"Cerebral vascular deposition of the amyloid-β (Aβ) peptide, a condition known as cerebral amyloid angiopathy (CAA), is associated with intracerebral hemorrhaging and contributes to disease progression in Alzheimer's disease (AD) and vascular cognitive impairment and dementia (VCID). Familial mutations at positions 22 and 23 within the Aβ peptide lead to early onset and severe CAA pathology. Here, we evaluate the effects of fibrillar Aβ peptides on the viability of primary-cultured human cerebral smooth muscle (HCSM) cells, which are the major site of amyloid deposition in cerebral blood vessel walls. Comparisons are made of the familial E22Q (Dutch) mutant of Aβ40 with wild-type Aβ40 and Aβ42. In agreement with previous studies, we find that there is a significant reduction in cell viability when Aβ40-Dutch or Aβ42-WT peptides are added to HCSM cell cultures as monomeric Aβ, whereas Aβ40-WT is relatively nontoxic. The binding of Aβ fibrils derived from sporadic CAA or familial Dutch-type CAA brain tissue to the membrane surface of HCSM cells does not result in a significant loss of cell viability. In contrast, when Aβ40-WT monomers and sporadic CAA fibrils are coincubated in HCSM cell cultures, there is a significant reduction in HCSM cell viability that is accompanied by an increase in cell surface fibril formation. Lastly, intrathecal administration of Aβ40-Dutch fibrillar seeds promotes fibrillar amyloid accumulation in the smooth muscle of meningeal vessels in the rTg-D transgenic rat model of CAA. Together, the present findings suggest that fibrillar Aβ seeds propagate the expansion of new amyloid fibrils on cerebral vascular smooth muscle, leading to membrane disruption and cell death.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":"64 5","pages":"1065-1078"},"PeriodicalIF":2.9,"publicationDate":"2025-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143539490","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Sortase-Mediated Fluorescent Labeling of eIF4E for Investigating Translation Initiation Mechanisms.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-03-04 Epub Date: 2025-02-19 DOI: 10.1021/acs.biochem.4c00851

Justin Pi, Simpson Joseph

引用次数: 0

Unraveling the Binding Mode of TSC2-Rheb through Protein Docking and Simulations.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-03-04 Epub Date: 2025-02-13 DOI: 10.1021/acs.biochem.4c00562

Berith F Pape, Shraddha Parate, Leif A Eriksson, Vibhu Jha

{"title":"Unraveling the Binding Mode of TSC2-Rheb through Protein Docking and Simulations.","authors":"Berith F Pape, Shraddha Parate, Leif A Eriksson, Vibhu Jha","doi":"10.1021/acs.biochem.4c00562","DOIUrl":"10.1021/acs.biochem.4c00562","url":null,"abstract":"Proteasome inhibitors (PIs) constitute the first line of therapy for multiple myeloma (MM). Despite the impressive clinical efficacy, MM remains fatal due to the development of drug resistance over time. During MM progression, stress responses to hypoxia and PIs suppress mammalian target of rapamycin complex 1 (mTORC1) activity by releasing tuberous sclerosis complex 2 (TSC2), which deactivates Ras homologue enriched in brain (Rheb), a crucial regulator of mTORC1. The efficacy of PIs targeting MM is enhanced when mTORC1 is hyperactivated. We thus propose that the inhibition of TSC2 will improve the efficacy of PIs targeting MM. To the best of our knowledge, no cocrystallized structure of the TSC2-Rheb complex has been reported. We therefore developed a representative model using the individual structures of TSC2 (PDB: 7DL2) and Rheb (PDB: 1XTS). Computational modeling involving an extensive protein-protein docking consensus approach was performed to determine the putative binding mode of TSC2-Rheb. The proposed docking poses were refined, clustered, and evaluated by MD simulations to explore the conformational dynamics and protein mobility, particularly at the drug-binding interface of TSC2-Rheb. Our results agree with the suggested binding mode of TSC2-Rheb previously reported in the literature. The results reported herein establish a basis for the development of new inhibitors blocking the binding of TSC2 and Rheb, aiming to reinstate mTORC1 activation and facilitate improved efficacy of PIs against multiple myeloma.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":"1006-1019"},"PeriodicalIF":2.9,"publicationDate":"2025-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11883811/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143412269","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Dynamics in the Phytophthora capsici Effector AVR3a11 Confirm the Core WY Domain Fold.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-03-04 Epub Date: 2025-02-20 DOI: 10.1021/acs.biochem.4c00660

James Tolchard, Vicki S Chambers, Laurence S Boutemy, Mark J Banfield, Tharin M A Blumenschein

{"title":"Dynamics in the Phytophthora capsici Effector AVR3a11 Confirm the Core WY Domain Fold.","authors":"James Tolchard, Vicki S Chambers, Laurence S Boutemy, Mark J Banfield, Tharin M A Blumenschein","doi":"10.1021/acs.biochem.4c00660","DOIUrl":"10.1021/acs.biochem.4c00660","url":null,"abstract":"Oomycete pathogens cause large economic losses in agriculture through diseases such as late blight (Phytophthora infestans), and stem and root rot of soybean (Phytophthora sojae). The effector protein AVR3a, from P. infestans, and its homologue AVR3a11 from Phytophthora capsici, are host-translocated effectors that interact with plant proteins to evade defense mechanisms and enable infection. Both proteins belong to the family of RXLR effectors and contain an N-terminal secretion signal, an RXLR motif for translocation into the host cell, and a C-terminal effector domain. Within this family, many proteins have been predicted to contain one or more WY domains as their effector domain, which is proposed to encompass a conserved minimal core fold containing three helices, further stabilized by additional helices or dimerization. In AVR3a11, a helical N-terminal extension to the core fold forms a four-helix bundle, as determined by X-ray crystallography. For a complete picture of the dynamics of AVR3a11, we have determined the solution structure of AVR3a11, and studied its dynamics in the fast time scale (ns-ps, from NMR relaxation parameters) and in the slow time scale (seconds to minutes, from hydrogen/deuterium exchange experiments). Hydrogen/deuterium exchange showed that the N-terminal helix is less stable than the other three helices, confirming the core fold originally proposed. Relaxation measurements confirm that AVR3a11 undergoes extensive conformational exchange, despite the uniform presence of fast motions in the spectral density function throughout most of its sequence. As functional residues are in the more mobile regions, flexibility in the slow/intermediate time scale may be functionally important.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":"1146-1156"},"PeriodicalIF":2.9,"publicationDate":"2025-03-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11883810/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143456263","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0