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Although biochemical experiments are the gold standard for determining protein functions, they are often time-consuming and labor-intensive. Here, we proposed a novel composite deep-learning method, Multi-source Knowledge Fusion for Gene Ontology prediction (MKFGO), to infer Gene Ontology (GO) attributes through integrating five complementary pipelines built on multi-source biological data. MKFGO was rigorously benchmarked on 1522 nonredundant proteins, demonstrating superior performance over 12 state-of-the-art function prediction methods. Comprehensive data analyses revealed that the major advantage of MKFGO lies in its two deep-learning components, handcrafted feature representation\u2013based GO prediction (HFRGO) and protein large language model (PLM)\u2013based GO prediction (PLMGO), which derive handcrafted features and PLM\u2013based features, respectively, from protein sequences in different biological views, with effective knowledge fusion at the decision-level. HFRGO leverages a long short-term memory (LSTM)\u2013attention network embedded with handcrafted features, in which the triplet loss\u2013based guilt-by-association strategy is designed to enhance the correlation between feature similarity and function similarity. PLMGO employs the PLM to capture feature embeddings with discriminative functional patterns from sequences. Meanwhile, another three components provide complementary insights for further improving prediction accuracy, driven by protein\u2013protein interaction, GO term probability, and protein-coding gene sequence, respectively. The source codes and models of MKFGO are freely available at https:\/\/github.com\/yiheng-zhu\/MKFGO.<\/jats:p>","DOI":"10.1093\/bib\/bbaf420","type":"journal-article","created":{"date-parts":[[2025,8,1]],"date-time":"2025-08-01T11:55:22Z","timestamp":1754049322000},"source":"Crossref","is-referenced-by-count":0,"title":["MKFGO: integrating multi-source knowledge fusion with pretrained language model for high-accuracy protein function prediction"],"prefix":"10.1093","volume":"26","author":[{"given":"Yi-Heng","family":"Zhu","sequence":"first","affiliation":[{"name":"College of Artificial Intelligence, Nanjing Agricultural University , 666 Binjiang Avenue, Jiangbei New District, Nanjing, Jiangsu Province, 211800 ,","place":["China"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Shuxin","family":"Zhu","sequence":"additional","affiliation":[{"name":"College of Artificial Intelligence, Nanjing Agricultural University , 666 Binjiang Avenue, Jiangbei New District, Nanjing, Jiangsu Province, 211800 ,","place":["China"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xuan","family":"Yu","sequence":"additional","affiliation":[{"name":"Department of Computer Science, City University of Hong Kong , 83 Tat Chee Avenue, Kowloon Tong, Hong Kong SAR (HKG), 999077 ,","place":["China"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"He","family":"Yan","sequence":"additional","affiliation":[{"name":"College of Information Science and Technology & Artificial Intelligence, Nanjing Forestry University , 159 Longpan Road, Xuanwu District, Nanjing, Jiangsu Province, 210037 ,","place":["China"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yan","family":"Liu","sequence":"additional","affiliation":[{"name":"School of Information Engineering, Yangzhou University , 196 Huayang West Road, Hanjiang District, Yangzhou, Jiangsu Province, 225000 ,","place":["China"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Xiaojun","family":"Xie","sequence":"additional","affiliation":[{"name":"College of Artificial Intelligence, Nanjing Agricultural University , 666 Binjiang Avenue, Jiangbei New District, Nanjing, Jiangsu Province, 211800 ,","place":["China"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6786-8053","authenticated-orcid":false,"given":"Dong-Jun","family":"Yu","sequence":"additional","affiliation":[{"name":"School of Computer Science and Engineering, Nanjing University of Science and Technology , 200 Xiaolingwei Street, Xuanwu District, Nanjing, Jiangsu Province, 210094 ,","place":["China"]}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Rui","family":"Ye","sequence":"additional","affiliation":[{"name":"College of Artificial Intelligence, Nanjing Agricultural University , 666 Binjiang Avenue, Jiangbei New District, Nanjing, Jiangsu Province, 211800 ,","place":["China"]}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"286","published-online":{"date-parts":[[2025,8,15]]},"reference":[{"key":"2025081501433620200_ref1","doi-asserted-by":"publisher","first-page":"823","DOI":"10.1038\/35015694","article-title":"Protein function in the post-genomic era","volume":"405","author":"Eisenberg","year":"2000","journal-title":"Nature"},{"key":"2025081501433620200_ref2","doi-asserted-by":"publisher","first-page":"6679","DOI":"10.1073\/pnas.0408930102","article-title":"Molecular dynamics and protein function","volume":"102","author":"Karplus","year":"2005","journal-title":"Proc Natl Acad Sci"},{"key":"2025081501433620200_ref3","doi-asserted-by":"publisher","first-page":"D330","DOI":"10.1093\/nar\/gky1055","article-title":"The gene ontology resource: 20 years and still GOing strong","volume":"47","author":"G. 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