Fusion tags and expression vector system for the expression of human parathyroid hormone (rhPTH)
Inventors
CHANDRASHEKARAN, Siddamadappa • HEMALATHA, Venkat
Assignees
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Publication Number
US-9580488-B2
Publication Date
2017-02-28
Expiration Date
Abstract
The present invention provides for fusion tags of 20 to 40 amino acids having a sequence as given in FIG. 1, fusion tags sharing homology with these fusion tags, or hybrid fusion tags comprising sequences of FIG. 1. The invention further provides an expression vector system comprising a fusion tag of the invention, and a process of obtaining recombinant human parathyroid hormone by using the fusion tags and the expression vector system of the invention.
Core Innovation
The invention relates to a recombinant fusion tag of 20-40 amino acid for expression and purification of recombinant human parathyroid hormone (rhPTH). The fusion tag comprises an amino acid sequence selected from the group consisting of amino acid sequences set forth in SEQ ID NO:1 through SEQ ID NO:7, and rhPTH includes rhPTH(1-34) as described in the provided content.
The system is an engineered expression vector system in E. coli that supports production of rhPTH using the defined short fusion tags. The expression vector design includes a His-tag and a TEV cleavage site positioned before rhPTH, together with the fusion tag and rhPTH to yield a fusion protein that can be processed to rhPTH.
The provided content further describes polynucleotides encoding the fusion tags and genetic constructs for expression. It also references obtaining rhPTH by synthesizing DNA encoding a 20-40 amino acid fusion tag, constructing and cloning a genetic construct, expressing a fusion protein in E. coli, then using TEV protease cleavage and partial purification.
Claims Coverage
The provided claim set centers on one independent claim defining a 20-40 amino-acid recombinant fusion tag, with additional dependent claim coverage for specific tag sequences, encoding polynucleotides, and expression vector and processing features. The independent claim set explicitly includes at least one independent claim: clm-00001.
Recombinant fusion tag of 20-40 amino acid
A recombinant fusion tag of 20-40 amino acid for expression and purification of recombinant human parathyroid hormone (rhPTH), said fusion tag comprising an amino acid sequence selected from the group consisting of SEQ ID NO:1 through SEQ ID NO:7.
Recombinant fusion tag sequence is SEQ ID NO.4
A recombinant fusion tag that comprises the amino acid sequence set forth in SEQ ID NO.4.
Polynucleotide encoding the recombinant fusion tag
A polynucleotide encoding the recombinant fusion tag described in claim 1.
Expression vector with His-tag, TEV cleavage site, and rhPTH
An expression vector wherein the fusion tag is fused with a His-tag, followed by a TEV cleavage site and rhPTH.
Expression vector having a pET28a vector backbone
An expression vector including a pET28a vector backbone.
Process for obtaining recombinant human parathyroid hormone (rhPTH)
A process for obtaining recombinant human parathyroid hormone (rhPTH) by synthesizing DNA encoding a 20-40 amino acid fusion tag, constructing and cloning a genetic construct, expressing a fusion protein in E. coli, then obtaining and partially purifying rhPTH.
Overall, the claim coverage protects specific short recombinant fusion tag sequences, coverage for a particular tag sequence, the encoding polynucleotide, and an expression vector architecture including His-tag and a TEV cleavage site arranged before rhPTH, optionally using a pET28a vector backbone, together with an E. coli expression and partial purification process to obtain rhPTH.
Stated Advantages
Not explicitly described in patent.
Documented Applications
Not explicitly described in patent.
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