Lactoferrin is an iron binding glycoprotein with multiple roles in the body. Its participation in apoptotic processes in cancer cells, its ability to modulate various reactions of the immune system, ...and its activity against a broad spectrum of pathogenic microorganisms, including respiratory viruses, have made it a protein of broad interest in pharmaceutical and food research and industry. In this review, we have focused on describing the most important functions of lactoferrin and the possible mechanisms of action that lead to its function.
Lactoferrin, a non-haeme iron-binding glycoprotein, known as the red fraction of milk, has been recently called a nutraceutical protein. Lactoferrin protein participates in some immunological ...processes and has an antimicrobial response against a broad spectrum of pathogens: it is also proposed to play a role in mechanisms of both neurodevelopment and neuropathy, and recently, the impact of lactoferrin against the metabolic syndrome was suggested. Other properties like anticancer, antioxidant and enzymatic activity have been reported. Because of this, the potential use of lactoferrin as a nutraceutical in the food industry makes it especially attractive to study and characterise. This review focuses on the description of lactoferrin as an important nutraceutical in science and food industry.
Breast cancer is the most frequently diagnosed cancer among women worldwide. Here, recombinant human lactoferrin (rhLf) expressed in
Pichia pastoris
was tested for its potential cytotoxic activity on ...a panel of six human breast cancer cell lines. The rhLf cytotoxic effect was determined via a live-cell HTS imaging assay. Also, confocal microscopy and flow cytometry protocols were employed to investigate the rhLf mode of action. The rhLf revealed an effective CC
50
of 91.4 and 109.46 µg/ml on non-metastatic and metastatic MDA-MB-231 cells, with favorable selective cytotoxicity index values, 11.68 and 13.99, respectively. Moreover, rhLf displayed satisfactory SCI values on four additional cell lines, MDA-MB-468, HCC70, MCF-7 and T-47D (1.55–3.34). Also, rhLf provoked plasma membrane blebbing, chromatin condensation and cell shrinkage in MDA-MB-231 cells, being all three apoptosis-related morphological changes. Also, rhLf was able to shrink the microfilaments, forming a punctuated cytoplasmic pattern in both the MDA-MB-231 and Hs-27 cells, as visualized in confocal photomicrographs. Moreover, performing flow cytometric analysis, rhLf provoked significant phosphatidylserine externalization, cell cycle arrest in the S phase and apoptosis-induced DNA fragmentation in MDA-MB-231 cells. Hence, rhLf possesses selective cytotoxicity on breast cancer cells. Also, rhLf caused apoptosis-associated morphologic changes, disruption of F-actin cytoskeleton organization, phosphatidylserine externalization, DNA fragmentation, and arrest of the cell cycle progression on triple-negative breast cancer MDA-MB-231 cells. Overall results suggest that rhLf is using the apoptosis pathway as its mechanism to inflict cell death. Findings warranty further evaluation of rhLf as a potential anti-breast cancer drug option.
Summary
Lactoferrin has gained extensive attention due to its ample biological properties. In this study, recombinant human lactoferrin carrying humanized glycosylation (rhLf-h-glycan) expressed in ...the yeast
Pichia pastoris
SuperMan5, which is genetically glycoengineered to efficiently produce functional humanized glycoproteins inclosing (Man)
5
(GlcNAc)
2
Asn-linked glycans, was analyzed, inspecting its potential toxicity against cancer cells. The live-cell differential nuclear staining assay was used to quantify the rhLf-h-glycan cytotoxicity, which was examined in four human cell lines: acute lymphoblastic leukemia (ALL) CCRF-CEM, T-cell lymphoblastic lymphoma SUP-T1, cervical adenocarcinoma HeLa, and as control, non-cancerous Hs27 cells. The defined CC
50
values of rhLf-h-glycan in CCRF-CEM, SUP-T1, HeLa, and Hs27 cells were 144.45 ± 4.44, 548.47 ± 64.41, 350 ± 14.82, and 3359.07 ± 164 µg/mL, respectively. The rhLf-h-glycan exhibited a favorable selective cytotoxicity index (SCI), preferentially killing cancer cells: 23.25 for CCRF-CEM, 9.59 for HeLa, and 6.12 for SUP-T1, as compared with Hs27 cells. Also, rhLf-h-glycan showed significant antiproliferative activity (
P
< 0.0001) at 24, 48, and 72 h of incubation on CCRF-CEM cells. Additionally, it was observed
via
fluorescent staining and confocal microscopy that rhLf-h-glycan elicited apoptosis-associated morphological changes, such as blebbing, nuclear fragmentation, chromatin condensation, and apoptotic bodies in ALL cells. Furthermore, rhLf-h-glycan-treated HeLa cells revealed shrinkage of the microfilament structures, generating a speckled/punctuated pattern and also caused PARP-1 cleavage, a hallmark of apoptosis. Moreover, in ALL cells, rhLf-h-glycan altered cell cycle progression inducing the G2/M phase arrest, and caused apoptotic DNA fragmentation. Overall, our findings revealed that rhLf-h-glycan has potential as an anticancer agent and therefore deserves further
in vivo
evaluation.
Maize is one of the most important crops worldwide and has been a target of nuclear-based transformation biotechnology to improve it and satisfy the food demand of the ever-growing global population. ...However, the maize plastid transformation has not been accomplished due to the recalcitrant condition of the crop.
In this study, we constructed two different vectors with homologous recombination sequences from maize (Zea mays var. LPC13) and grass (Bouteloua gracilis var. ex Steud) (pZmcpGFP and pBgcpGFP, respectively). Both vectors were designed to integrate into rrn23S/rrn16S from an inverted repeat region in the chloroplast genome. Moreover, the vector had the mgfp5 gene driven by Prrn, a leader sequence of the atpB gene and a terminator sequence from the rbcL gene. Also, constructs have an hph gene as a selection marker gene driven by Prrn, a leader sequence from rbcL gene and a terminator sequence from the rbcL gene. Explants of maize, tobacco and Escherichia coli cells were transformed with both vectors to evaluate the transitory expression–an exhibition of green and red fluorescent light under epifluorescence microscopy. These results showed that both vectors were expressed; the reporter gene in all three organisms confirmed the capacity of the vectors to express genes in the cell compartments.
This paper is the first report of transient expression of GFP in maize embryos and offers new information for genetically improving recalcitrant crops; it also opens new possibilities for the improvement in maize chloroplast transformation with these vectors.
How to cite: Arévalo-Gallegos S, Varela-Rodríguez H, Lugo-Aguilar H, et al. Transient expression of a green fluorescent protein in tobacco and maize chloroplast. Electron J Biotechnol 2020;44. https://doi.org/10.1016/j.ejbt.2020.01.008
Lactoferrin a multiple bioactive protein: An overview García-Montoya, Isui Abril; Cendón, Tania Siqueiros; Arévalo-Gallegos, Sigifredo ...
Biochimica et biophysica acta,
03/2012, Letnik:
1820, Številka:
3
Journal Article
Recenzirano
Odprti dostop
Lactoferrin (Lf) is an 80kDa iron-binding glycoprotein of the transferrin family. It is abundant in milk and in most biological fluids and is a cell-secreted molecule that bridges innate and adaptive ...immune function in mammals. Its protective effects range from anticancer, anti-inflammatory and immune modulator activities to antimicrobial activities against a large number of microorganisms. This wide range of activities is made possible by mechanisms of action involving not only the capacity of Lf to bind iron but also interactions of Lf with molecular and cellular components of both hosts and pathogens.
This review summarizes the activities of Lf, its regulation and potential applications.
The extensive uses of Lf in the treatment of various infectious diseases in animals and humans has been the driving force in Lf research however, a lot of work is required to obtain a better understanding of its activity.
The large potential applications of Lf have led scientists to develop this nutraceutical protein for use in feed, food and pharmaceutical applications. This article is part of a Special Issue entitled Molecular Mechanisms of Iron Transport and Disorders.
► It is well known that the mechanisms of action of antimicrobial activity are not fully understood. ► Important advances were made in heterologous Lf expression in prokaryotic systems. ► In the near future we will see the exploitation of the full capacity of lactoferrin expression in plants.
Immunomodulatory effects of lactoferrin Tania SIQUEIROS-CENDON Sigifredo AREVALO-GALLEGOS Blanca Flor IGLESIAS-FIGUEROA Isui Abril GARCiA-MONTOYA Jose SALAZAR-MARTIN EZ Quintin RASCON-CRUZ
Acta pharmacologica Sinica,
05/2014, Letnik:
35, Številka:
5
Journal Article
Recenzirano
Odprti dostop
Lactoferrin （Lf） is an iron-binding glycoprotein of the transferrin family, which is expressed in most biological fluids with particularly high levels in mammalian milk. Its multiple activities lie ...in its capacity to bind iron and to interact with the molecular and cellular components of hosts and pathogens. Lf can bind and sequester lipopolysaccharides, thus preventing pro-inflammatory pathway activation, sepsis and tissue damages. Lf is also considered a cell-secreted mediator that bridges the innate and adaptive immune responses. In the recent years much has been learned about the mechanisms by which Lf exerts its activities. This review summarizes the recent advances in understanding the mechanisms underlying the multifunctional roles of Lf, and provides a future perspective on its potential prophylactic and therapeutic applications.
Due to their ability to produce isomaltulose, sucrose isomerases are enzymes that have caught the attention of researchers and entrepreneurs since the 1950s. However, their low activity and stability ...at temperatures above 40 °C have been a bottleneck for their industrial application. Specifically, the instability of these enzymes has been a challenge when it comes to their use for the synthesis and manufacturing of chemicals on a practical scale. This is because industrial processes often require biocatalysts that can withstand harsh reaction conditions, like high temperatures. Since the 1980s, there have been significant advancements in the thermal stabilization engineering of enzymes. Based on the literature from the past few decades and the latest achievements in protein engineering, this article systematically describes the strategies used to enhance the thermal stability of sucrose isomerases. Additionally, from a theoretical perspective, we discuss other potential mechanisms that could be used for this purpose.
In this study, bovine lactoferrin (bLf), an iron-binding glycoprotein considered an important nutraceutical protein because of its several properties, was expressed in Pichia pastoris KM71-H under ...AOX1 promoter control, using pJ902 as the recombinant plasmid. Dot blotting analysis revealed the expression of recombinant bovine lactoferrin (rbLf) in Pichia pastoris. After Bach fermentation and purification by molecular exclusion, we obtained an expression yield of 3.5 g/L of rbLf. rbLf and predominantly pepsin-digested rbLf (rbLfcin) demonstrated antibacterial activity against Escherichia coli (E. coli) BL21DE3, Staphylococcus aureus (S. aureus) FRI137, and, in a smaller percentage, Pseudomonas aeruginosa (Ps. Aeruginosa) ATCC 27833. The successful expression and characterization of functional rbLf expressed in Pichia pastoris opens a prospect for the development of natural antimicrobial agents produced recombinantly.
Chloroplast biotechnology has emerged as a promissory platform for the development of modified plants to express products aimed mainly at the pharmaceutical, agricultural, and energy industries. This ...technology’s high value is due to its high capacity for the mass production of proteins. Moreover, the interest in chloroplasts has increased because of the possibility of expressing multiple genes in a single transformation event without the risk of epigenetic effects. Although this technology solves several problems caused by nuclear genetic engineering, such as turning plants into safe bio-factories, some issues must still be addressed in relation to the optimization of regulatory regions for efficient gene expression, cereal transformation, gene expression in non-green tissues, and low transformation efficiency. In this article, we provide information on the transformation of plastids and discuss the most recent achievements in chloroplast bioengineering and its impact on the biopharmaceutical and agricultural industries; we also discuss new tools that can be used to solve current challenges for their successful establishment in recalcitrant crops such as monocots.
How to cite: Quintín Rascón-Cruz Q, González-Barriga CD, Iglesias-Figueroa BF, et al. Plastid transformation: Advances and challenges for its implementation in agricultural crops. Electron J Biotechnol 2021;51. https://doi.org/10.1016/j.ejbt.2021.03.005
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