fusion partner
Recently Published Documents


TOTAL DOCUMENTS

490
(FIVE YEARS 131)

H-INDEX

49
(FIVE YEARS 4)

2022 ◽  
Vol 27 ◽  
pp. 300582
Author(s):  
Rofieda R. Alwaqfi ◽  
Jonathan J. Davick ◽  
Aaron D. Bossler
Keyword(s):  

2022 ◽  
Vol 189 ◽  
pp. 105991
Author(s):  
Sreejith Raran-Kurussi ◽  
Sarawata B. Sharwanlal ◽  
Deepa Balasubramanian ◽  
Kaustubh R. Mote

AMB Express ◽  
2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Takashi Koyanagi ◽  
Ayumi Hara ◽  
Kanako Kobayashi ◽  
Yuji Habara ◽  
Akira Nakagawa ◽  
...  

AbstractPeptidyl-prolyl cis-trans isomerase (PPIase, EC 5.2.1.8) catalyzes the racemization reaction of proline residues on a polypeptide chain. This enzyme is also known to function as a molecular chaperon to stabilize protein conformation during the folding process. In this study, we noted FK506 binding protein (FKBP)-type PPIase from a hyperthemophilic archaeon Thermococcus sp. strain KS-1 (PPIase KS−1) to improve the solubility of Pseudomonas putida aromatic amino acid decarboxylase (AADC) that is an indispensable enzyme for fermentative production of plant isoquinoline alkaloids. AADC fused N-terminally with the PPIase KS−1 (PPIase KS−1-AADC), which was synthesized utilizing Escherichia coli host, showed improved solubility and, consequently, the cell-free extract from the recombinant strain exhibited 2.6- to 3.4-fold elevated AADC activity than that from the control strain that expressed the AADC gene without PPIase KS−1. On the other hand, its thermostability was slightly decreased by fusing PPIase KS−1. The recombinant E. coli cells expressing the PPIase KS−1-AADC gene produced dopamine and phenylethylamine from L-dopa and phenylalanine by two- and threefold faster, respectively, as compared with the control strain. We further demonstrated that the efficacy of PPIase KS−1-AADC in solubility and activity enhancement was a little but obviously higher than that of AADC fused N-terminally with NusA protein, which has been assumed to be the most effective protein solubilizer. These results suggest that PPIase KS−1 can be used as one of the best choices for producing heterologous proteins as active forms in E. coli.


2021 ◽  
Vol 22 (23) ◽  
pp. 12906
Author(s):  
Masaya Mitsumoto ◽  
Kanna Sugaya ◽  
Kazuki Kazama ◽  
Ryosuke Nakano ◽  
Takahiro Kosugi ◽  
...  

G-protein coupled receptors (GPCRs) are known for their low stability and large conformational changes upon transitions between multiple states. A widely used method for stabilizing these receptors is to make chimeric receptors by fusing soluble proteins (i.e., fusion partner proteins) into the intracellular loop 3 (ICL3) connecting the transmembrane helices 5 and 6 (TM5 and TM6). However, this fusion approach requires experimental trial and error to identify appropriate soluble proteins, residue positions, and linker lengths for making the fusion. Moreover, this approach has not provided state-targeting stabilization of GPCRs. Here, to rationally stabilize a class A GPCR, adenosine A2A receptor (A2AR) in a target state, we carried out the custom-made de novo design of α-helical fusion partner proteins, which can fix the conformation of TM5 and TM6 to that in an inactive state of A2AR through straight helical connections without any kinks or intervening loops. The chimeric A2AR fused with one of the designs (FiX1) exhibited increased thermal stability. Moreover, compared with the wild type, the binding affinity of the chimera against the agonist NECA was significantly decreased, whereas that against the inverse agonist ZM241385 was similar, indicating that the inactive state was selectively stabilized. Our strategy contributes to the rational state-targeting stabilization of GPCRs.


2021 ◽  
pp. 106689692110479
Author(s):  
Gorana Gasljevic ◽  
Matthias S. Matter ◽  
Olga Blatnik ◽  
Mojca Unk ◽  
Stefan Dirnhofer

Background: NUT carcinoma is a highly aggressive and rare subset of squamous cell carcinoma with grim prognosis. It is under-recognized by both pathologists and oncologists. Recognition is challenging due to its rareness and the fact that its clinical and laboratory features as well as morphological and immunohistochemical characteristics may mimic other malignancies. Case presentation: An interesting case of NUT carcinoma in a 47-year-old male with a large tumor mass in the inferior part of the mediastinum and left lung and increased levels of serum alpha fetoprotein (AFP) is described. Immunohistochemical analysis of both the primary tumor in a bronchoscopy specimen and an excisional biopsy of a subcutaneous metastasis showed positivity for AFP and leukocyte common antigen (LCA) that were misleading and resulted in diagnostic pitfalls of mediastinal germ cell tumor (clinically) and hematolymphoid neoplasm (pathologic report). Immunohistochemical demonstration of NUT protein expression revealed the proper diagnosis, which was further confirmed by RNA sequencing revealing a BRD4- NUTM1 gene fusion.Conclusions: Since NUT carcinoma can show a wide spectrum of histological and immunophenotypic features and can clinically mimic other tumors, use of RNA sequencing with identification of specific NUTM1 fusion partner could be crucial when there are discrepant clinical and histopathological findings. As well, since the category of so-called NUTM1-rearranged neoplasms is rapidly expanding, identification of NUTM1 fusion partner may be essential for the appropriate clinical management.


Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 3480-3480
Author(s):  
Irina Pushel ◽  
Midhat S. Farooqi ◽  
Byunggil Yoo ◽  
Daniel Louiselle ◽  
Tomi Pastinen ◽  
...  

Abstract Introduction: Infant acute lymphoblastic leukemia (ALL) is an aggressive subtype of leukemia with low rates of survival. Rearrangements involving the KMT2A gene are associated with poor prognosis for infants with this cancer. Although many patients with KMT2A rearrangements (KMT2A-r) ultimately relapse, some do not. The molecular basis for this distinction has not yet been determined. There is evidence to suggest that cancers with distinct KMT2A fusion partners show distinct patterns of gene expression, though these differences have not previously been linked to prognostic outcomes. Here we utilize single-cell genomic analysis for infant ALL samples with and without KMT2A-r taken at diagnosis to explore transcriptional dynamics and identify gene expression programs with potential prognostic value. Methods: We performed 10x Chromium single-cell multiome sequencing to obtain both gene expression (scRNA-seq) and chromatin accessibility (scATAC-seq) data for blood and/or bone marrow samples obtained from a total of 34 infant ALL patients at time of diagnosis. Of these, 19 KMT2A-r patients later relapsed, 6 KMT2A-r patients did not relapse, and 9 patients did not show KMT2A rearrangement. Sequencing data were processed and aggregated with cellranger-arc, with normalization and differential gene expression performed using the Seurat package for gene expression analysis. Differential gene expression was performed between the three groups described above, (KMT2A-r + relapse, KMT2A-r no relapse, no KMT2A-r), then further subdivided for KMT2A-r cases based on the KMT2A partner gene (MLLT1 n=13 or AFF1 n=12). Results: In preliminary scRNA- and scATAC-seq in infant ALL patients, we had observed that cancer cells from individual patients tend to cluster separately, while non-blast cell populations showed similar interindividual patterns, suggesting a high degree of divergence among blast cell transcriptional programs. We replicated this finding here in larger patient samples, demonstrating no overt similarity based on KMT2A-r status or whether patients later relapsed. Strikingly, samples did broadly cluster according to KMT2A rearrangement partner gene, indicating that this appears to be a major driver of the transcriptional program in infant ALL patients. Differential expression analysis within the KMT2A-MLLT1 and KMT2A-AFF1 samples separately revealed expression of other genes associated with particular KMT2A fusion genes. Specifically, we observed that expression of HOXA genes was mostly restricted to KMT2A-MLLT1 samples. Consistent with previous results, we observed that within KMT2A-AFF1 samples there were two distinct groups of cells with mutually exclusive expression of HOXA9 or IRX1. Furthermore, the limited HOX gene expression observed in KMT2A-AFF1 samples was enriched in patients who did not relapse, supporting previous observations that the KMT2A-AFF1 samples expressing IRX1 comprise a more aggressive leukemia. Conclusions: Utilizing a single-cell transcriptomic approach enabled us to identify gene expression programs associated with good and poor prognosis in KMT2A-r infant ALL cases. We found that the KMT2A fusion partner gene appears to drive a large portion of the transcriptional heterogeneity observed across KMT2A-r samples. By treating the KMT2A-MLLT1 and KMT2A-AFF1 samples separately and exploring transcriptional dynamics within these groups, we identified transcriptional differences between patients who did and did not relapse in the presence of a KMT2A-AFF1 fusion. We are continuing to explore these data to identify prognostic markers in KMT2A-MLLT1 patients. Figure 1. Mutually exclusive gene expression programs distinguish subsets of infant ALL samples. A) Uniform manifold approximation and projection (UMAP) visualization of gene expression in individual cells across all patient samples at diagnosis, each color corresponding to a single patient (n=34). B) Cells colored according to KMT2A-r status and relapse or lack thereof. C) Cells colored according to KMT2A fusion partner gene. D) HOXA9 expression among KMT2A-AFF1 patients is strongly biased to patients that do not relapse. E) In KMT2A-AFF1 samples, HOXA9 and IRX1 show mutually exclusive expression patterns. Figure 1 Figure 1. Disclosures Brown: Novartis: Membership on an entity's Board of Directors or advisory committees; Takeda: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees; Kura: Membership on an entity's Board of Directors or advisory committees; KIte: Membership on an entity's Board of Directors or advisory committees.


Blood ◽  
2021 ◽  
Vol 138 (Supplement 1) ◽  
pp. 3430-3430
Author(s):  
Brian Ball ◽  
Shukaib Arslan ◽  
Paul Koller ◽  
Joyce Murata-Collins ◽  
Michelle Afkhami ◽  
...  

Abstract Background Rearrangements of lysine methyltransferase 2A (KMT2A) gene, previously known as the MLL gene, occur in 3 to 5% of adult patients with de novo AML and are enriched in therapy-related disease after treatment with topoisomerase II inhibitors (Bill, M et al. PNAS. 2020). KMT2A encodes a histone H3 lysine 4 methyltransferase and KMT2A rearrangements result in fusion proteins that induce aberrant Hox gene expression. (Armstrong, SA et al. Nature. 2002) Among patients with KMT2A rearranged AML (rAML) receiving intensive chemotherapy, the fusion partner impacts prognosis, and KMT2A-MLLT3 is associated with an intermediate risk while other KMT2A rearrangements are associated with adverse risk in the ELN classification. (Dohner, H et al. Blood. 2017) Here we evaluate the outcomes of patients with newly diagnosed and relapsed or refractory (R/R) AML with KMT2A rearrangements receiving venetoclax and hypomethylating agents (HMA). Methods Medical records of 333 patients with newly diagnosed or R/R AML receiving venetoclax in combination with HMAs at City of Hope National Medical Center between 11/1/2015 and 4/15/2020 were reviewed. Criteria for inclusion were a pathologically confirmed diagnosis of AML, age > 18 years, treatment with either decitabine or azacitidine in combination with venetoclax. KMT2A rearrangements were detected by karyotype and confirmed by FISH or RNA sequencing. Responses were evaluated per the ELN criteria (Dohner, H et al. Blood. 2017) Minimal residual disease flow cytometry was performed at the University of Washington. Patient characteristics were summarized by frequency and associations between overall response and patient and disease characteristics were tested by Fisher's exact test. OS was evaluated by the Kaplan-Meier method and the difference between groups was determined by log-rank test. All statistical analyses were performed using SPSS and Prism. Results We identified 18 patients (5.4%) with KMT2A rAML who met criteria for inclusion. MLLT3 was the predominant fusion partner, occurring in nine patients followed by ELL (n=2), AFDN (n=2), MLLT6 (n=1), MLLT10 (n=1), AFF1 (n=1), CBL (n=1), and TET1 (n=1). The cohort included both newly diagnosed (n=10) and R/R (n=8) AML patients. 44% had therapy-related or secondary AML. NRAS or KRAS mutations occurred in 4 out of 13 patients (31%) with available sequencing prior to treatment. Decitabine was the predominant HMA used in combination with venetoclax and 56% of all patients received 10-day dosing during the first cycle. For the total cohort, 9 patients achieved an overall response (ORR 50%), including 8 patients with a complete remission (CR/CRi 44%) and 1 (6%) patient with a morphologic leukemia free state, Figure 1. All six of the responders who were tested for MRD were negative. For treatment naïve patients, we observed a CR/CRi rate of 70% and a median survival of 11 months. On univariate analysis, R/R disease was the only factor associated with a significant decrease in response (ORR 12.5% vs. 80%, p=0.015, Table 1). With a median follow-up of 14.4 months for responding patients, median OS for the cohort was 6.59 months and 19.15 months for responding patients (Figure 2). The presence of NRAS or KRAS mutations was the only factor significantly impacting survival (HR 6.04, log rank 0.05, Table 1). Notably, the KMT2A fusion partner type did not impact response or survival. Allogeneic stem cell transplant was performed in 4 out of 9 (44%) responding patients. Conclusion Here, we show that venetoclax in combination with HMA led to a high rate of response and prolonged survival in a high-risk KMT2A rAML population. The outcomes of newly diagnosed KMT2A rAML patients after treatment with venetoclax and HMA in this study are similar to chemotherapy outcomes in patients aged < 60 years (CR 68% and median OS 0.9 months; Bill, M et al. PNAS. 2020). Consistent with previous studies, we found that MLLT3 was the most common fusion partner, occurring in 50% of patients and that RAS mutations were also common (~30%). In contrast to what has been reported for chemotherapy outcomes and in the ELN classification, the KMT2A-MLLT3 translocation was not associated with improved outcomes when compared to other KMT2A translocations. While this study was limited in being retrospective and having a small and heterogeneous population, our findings suggest that venetoclax and HMA are effective in KMT2A rAML and warrant further investigation. Figure 1 Figure 1. Disclosures Koller: Novartis: Consultancy. Al Malki: Hansa Biopharma: Consultancy; Rigel Pharma: Consultancy; Neximmune: Consultancy; Jazz Pharmaceuticals, Inc.: Consultancy; CareDx: Consultancy. Aribi: Seagen: Consultancy. Ali: Incyte: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; BMS: Speakers Bureau; CTI BioPharma: Membership on an entity's Board of Directors or advisory committees. Marcucci: Novartis: Other: Speaker and advisory scientific board meetings; Agios: Other: Speaker and advisory scientific board meetings; Abbvie: Other: Speaker and advisory scientific board meetings. Pullarkat: AbbVie, Amgen, Genentech, Jazz Pharmaceuticals, Novartis, Pfizer, and Servier: Membership on an entity's Board of Directors or advisory committees; Amgen, Dova, and Novartis: Consultancy, Honoraria.


2021 ◽  
pp. 106689692110498
Author(s):  
Haider Mejbel ◽  
Gene P. Siegal ◽  
Shi Wei

Tenosynovial giant cell tumors typically arise in the synovium of joints, bursae, or tendon sheaths. They may occur in an intra- or extra-articular location and can be divided into localized and diffuse types. The neoplastic nature of the lesion has been supported by a recurrent CSF1 gene rearrangement in a small subset of lesional cells, of which the most common fusion partner is COL6A3. Herein, we report a case of intramuscular localized tenosynovial giant cell tumor harboring a novel CSF1-CD96 fusion transcript, thus expanding the molecular profile of this tumor.


Author(s):  
Murad Mollaev ◽  
Artur Zabolotskii ◽  
Neonila Gorokhovets ◽  
Elena Nikolskaya ◽  
Maria Sokol ◽  
...  

Abstract Background Difficult to express peptides are usually produced by co-expression with fusion partners. In this case, a significant mass part of the recombinant product falls on the subsequently removed fusion partner. On the other hand, multimerization of peptides is known to improve its proteolytic stability in E. coli due to the inclusion of body formation, which is sequence specific. Thereby, the peptide itself may serve as a fusion partner and one may produce more than one mole of the desired product per mole of fusion protein. This paper proposes a method for multimeric production of a human alpha-fetoprotein fragment with optimized multimer design and processing. This fragment may further find its application in the cytotoxic drug delivery field or as an inhibitor of endogenous alpha-fetoprotein. Results Multimerization of the extended alpha-fetoprotein receptor-binding peptide improved its stability in E. coli, and pentamer was found to be the largest stable with the highest expression level. As high as 10 aspartate-proline bonds used to separate peptide repeats were easily hydrolyzed in optimized formic acid-based conditions with 100% multimer conversion. The major product was represented by unaltered functional alpha-fetoprotein fragment while most side-products were its formyl-Pro, formyl-Tyr, and formyl-Lys derivatives. Single-step semi-preparative RP-HPLC was enough to separate unaltered peptide from the hydrolysis mixture. Conclusions A recombinant peptide derived from human alpha-fetoprotein can be produced via multimerization with subsequent formic acid hydrolysis and RP-HPLC purification. The reported procedure is characterized by the lower reagent cost in comparison with enzymatic hydrolysis of peptide fusions and solid-phase synthesis. This method may be adopted for different peptide expression, especially with low amino and hydroxy side chain content.


2021 ◽  
Author(s):  
Xuefeng Li ◽  
Baorong Zhang ◽  
Quan Hu ◽  
Changchao Chen ◽  
Lu Liu ◽  
...  

Abstract The methods developed for efficient insoluble protein production are less well explored. Our data demonstrated that PagP, an E. coli outer membrane protein with high β-sheet content, could function as an efficient fusion partner for inclusion body-targeted expression of antimicrobial peptide Magainin II, Metchnikowin and Andropin. The primary structure of a given polypeptide determines to a large extent its propensity to aggregate. The aggregation “hot spots” (HSs) in PagP was subsequently analyzed with the web-based software AGGRESCAN, leading to identification of the C-terminal region with high dense distribution of HSs. The absolute yields of recombinant antimicrobial peptide Metchnikowin and Andropin could be increased significantly when expressed in fusion with this version of PagP. Moreover, a Proline-rich region was found in the β-strands of PagP. Substitution for these prolines by residues with high β-sheet propensity and hydrophobicity significantly improved its ability to form aggregates, and greatly increased the yield of the recombinant passenger peptides. Fewer examples have been presented to separate the recombinant target proteins expressed in fusion inclusion bodies. Here, we reported an artificial linker peptide NHT with three motifs, by which separation and purification of the authentic recombinant antimicrobial peptides could be implemented.


Sign in / Sign up

Export Citation Format

Share Document