Characterization of methanol utilization negative
            Pichia pastoris
            for secreted protein production: New cultivation strategies for current and future applications

Abstract Caprine chorion, allantois and amnion from days 23, 28, 35, 39 and 45, and yolk sac from day 23 of pregnancy were isolated by dissection and cultured for 24 h in modified minimum essential medium in the presence of [35S] methionine. De novo-synthesized proteins released into the culture medium were analyzed by two-dimensional PAGE and fluorography. Patterns of protein production by these isolated extraembryonic membranes remained relatively unchanged from days 23 to 45 of pregnancy. Electrophoretic profiles of proteins synthesized by allantois and amnion were identical but distinct from that produced by chorion. Yolk sac was the major source of serum-like proteins. An acidic (pI 5·3–6·3) 22 kDa protein, which consisted of four isoelectric variants, was produced by all extraembryonic membranes and demonstrated to immunoreact with antiserum produced against bovine placental retinol-binding protein (RBP). Limited N-terminal sequence analysis of one major isoform indicated that the protein had complete homology with bovine RBP over the first 15 amino acids. Immunoreactive RBP was localized in epithelial cells lining the chorion, allantois and amnion. In this study, we have characterized and compared protein production by isolated extraembryonic membranes through days 23 to 45 of pregnancy and identified the 22 kDa protein as caprine RBP of placental origin. Journal of Endocrinology (1995) 146, 527–534

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Purification and characterization of asparaginase II from Saccharomyces cerevisiae cloned in Pichia pastoris: a study on a possible antileukemic drug

BMC Proceedings ◽

10.1186/1753-6561-8-s4-p78 ◽

2014 ◽

Vol 8 (S4) ◽

Cited By ~ 1

Author(s):

Luciana Facchinetti de Castro Girão ◽

Surza Lucia Gonçalves da Rocha ◽

Ricardo Sobral Teixeira ◽

Maria Antonieta Ferrara ◽

Jonas Perales ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Pichia Pastoris ◽

Purification And Characterization

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Heterologous expression in Pichia pastoris and biochemical characterization of the unmodified sulfatase from Fusarium proliferatum LE1

Protein Engineering Design and Selection ◽

10.1093/protein/gzx047 ◽

2017 ◽

Vol 30 (8) ◽

pp. 571-571 ◽

Cited By ~ 1

Author(s):

Svetlana A Korban ◽

Kirill S Bobrov ◽

Maria A Maynskova ◽

Stanislav N Naryzhny ◽

Olga L Vlasova ◽

...

Keyword(s):

Pichia Pastoris ◽

Heterologous Expression ◽

Biochemical Characterization ◽

Fusarium Proliferatum

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Protein expression and extraction of hard-to-produce proteins in the periplasmic space of Escherichia coli v1

10.17504/protocols.io.bxxxpppn ◽

2021 ◽

Author(s):

Cristina Hernandez Rollan ◽

Kristoffer Bach Falkenberg ◽

Maja Rennig ◽

Andreas Birk Bertelsen ◽

Morten Norholm

Keyword(s):

Protein Production ◽

Expression System ◽

Gram Negative Bacteria ◽

Gram Negative ◽

E Coli ◽

New Family ◽

Lytic Polysaccharide Monooxygenases ◽

Strain Bl21 ◽

Polysaccharide Monooxygenases

E. coli is a gram-negative bacteria used mainly in academia and in some industrial scenarios, as a protein production workhorse. This is due to its ease of manipulation and the range of genetic tools available. This protocol describes how to express proteins in the periplasm E. coli with the strain BL21 (DE3) using a T7 expression system. Specifically, it describes a series of steps and tips to express "hard-to-express" proteins in E. coli, as for instance, LPMOs. The protocol is adapted from Hemsworth, G. R., Henrissat, B., Davies, G. J., and Walton, P. H. (2014) Discovery and characterization of a new family of lytic polysaccharide monooxygenases. Nat. Chem. Biol.10, 122–126. .

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