{"doi":"10.1101/2023.01.23.525242","title":"Direct Tests of Cytochrome Function in the Electron Transport Chain of Malaria Parasites","abstract":"ABSTRACT The mitochondrial electron transport chain (ETC) of Plasmodium malaria parasites is a major antimalarial drug target, but critical cytochrome functions remain unstudied and enigmatic. Parasites express two distinct cyt c homologs ( c and c -2) with unusually sparse sequence identity and uncertain fitness contributions. P. falciparum cyt c -2 is the most divergent eukaryotic cyt c homolog currently known and has sequence features predicted to be incompatible with canonical ETC function. We tagged both cyt c homologs and the related cyt c 1 for inducible knockdown. Translational repression of cyt c and cyt c 1 was lethal to parasites, which died from ETC dysfunction and impaired ubiquinone recycling. In contrast, cyt c -2 knockdown or knock-out had little impact on blood-stage growth, indicating that parasites rely fully on the more conserved cyt c for ETC function. Biochemical and structural studies revealed that both cyt c and c -2 are hemylated by holocytochrome c synthase, but UV-vis absorbance and EPR spectra strongly suggest that cyt c -2 has an unusually open active site in which heme is stably coordinated by only a single axial amino-acid ligand and can bind exogenous small molecules. These studies provide a direct dissection of cytochrome functions in the ETC of malaria parasites and identify a highly divergent Plasmodium cytochrome c with molecular adaptations that defy a conserved role in eukaryotic evolution. SIGNIFICANCE STATEMENT Mitochondria are critical organelles in eukaryotic cells that drive oxidative metabolism. The mitochondrion of Plasmodium malaria parasites is a major drug target that has many differences from human cells and remains poorly studied. One key difference from humans is that malaria parasites express two cytochrome c proteins that differ significantly from each other and play untested and uncertain roles in the mitochondrial electron transport chain (ETC). Our study revealed that one cyt c is essential for ETC function and parasite viability while the second, more divergent protein has unusual structural and biochemical properties and is not required for growth of blood-stage parasites. This work elucidates key biochemical properties and evolutionary differences in the mitochondrial ETC of malaria parasites.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2023,"id":402566,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":0,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9509,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2023-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1063004,"name":"Henry Wienkers","orcid":"0000-0001-7948-3241","position":1,"is_corresponding":false},{"id":1063329,"name":"Rebecca G. Marvin","orcid":null,"position":2,"is_corresponding":false},{"id":1063005,"name":"Shai-anne Nalder","orcid":"0000-0002-6892-9321","position":3,"is_corresponding":false},{"id":1063006,"name":"Aldo E. García-Guerrero","orcid":"0000-0001-6751-8304","position":4,"is_corresponding":false},{"id":1063007,"name":"Peter E. VanNatta","orcid":"0000-0003-1360-579X","position":5,"is_corresponding":false},{"id":344710,"name":"Yasaman Jami‐Alahmadi","orcid":"0000-0001-8289-2222","position":6,"is_corresponding":false},{"id":1063008,"name":"Amanda Mixon Blackwell","orcid":"0000-0002-7473-5822","position":7,"is_corresponding":false},{"id":578665,"name":"Frank G. Whitby","orcid":"0000-0003-3511-2216","position":8,"is_corresponding":false},{"id":14958,"name":"James A. Wohlschlegel","orcid":"0000-0003-3399-901X","position":9,"is_corresponding":false},{"id":675303,"name":"Matthew T. Kieber‐Emmons","orcid":"0000-0002-6357-5579","position":10,"is_corresponding":false},{"id":666493,"name":"Christopher P. Hill","orcid":"0000-0001-6796-7740","position":11,"is_corresponding":false},{"id":676585,"name":"Paul A. Sigala","orcid":"0000-0002-3464-3042","position":12,"is_corresponding":false},{"id":1063003,"name":"Tanya J. Espino-Sanchez","orcid":"0000-0002-7076-2145","position":0,"is_corresponding":true}],"reference_count":97,"raw_metadata":{"citation_network_status":"fetched"},"created_at":"2026-07-19T01:20:27.903246Z","pmid":"36747727","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}