A method for [3H]mannose labeling of Asn-linked oligosaccharides on recombinant glycoproteins synthesized in Xenopus oocytes

Alan B. Cantor; Stuart Kornfeld

doi:10.1016/0003-2697(92)90427-9

A method for [³H]mannose labeling of Asn-linked oligosaccharides on recombinant glycoproteins synthesized in Xenopus oocytes

Alan B. Cantor
, Stuart Kornfeld

Research output: Contribution to journal › Article › peer-review

2 Scopus citations

Abstract

We have developed an efficient method for labeling the Asn-linked oligosaccharides of recombinant glycoproteins synthesized in Xenopus laevis oocytes. By coinjecting GDP-[3,4-³H]mannose with mRNA for human cathepsin D, it was possible to incorporate as much as 1800 cpm per oocyte into each of the two Asn-linked oligosaccharides of this glycoprotein. Overall, about 50% of the microinjected GDP-[3,4-³H]mannose was incorporated into Asn-linked oligosaccharides, a 10-fold greater value than that obtained when [2-³H]mannose was microinjected. Less than 10% of the injected GDP-[3,4-³H]mannose was metabolized to water or converted to amino acids. This technique should facilitate studies of Asn-linked oligosaccharide biosynthesis, processing, and structure in recombinant proteins synthesized in Xenopus oocytes.

Original language	English
Pages (from-to)	220-226
Number of pages	7
Journal	Analytical Biochemistry
Volume	205
Issue number	2
DOIs	https://doi.org/10.1016/0003-2697(92)90427-9
State	Published - Sep 1992

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@article{125e4295ed3f4b0089130de9b06b1193,

title = "A method for [3H]mannose labeling of Asn-linked oligosaccharides on recombinant glycoproteins synthesized in Xenopus oocytes",

abstract = "We have developed an efficient method for labeling the Asn-linked oligosaccharides of recombinant glycoproteins synthesized in Xenopus laevis oocytes. By coinjecting GDP-[3,4-3H]mannose with mRNA for human cathepsin D, it was possible to incorporate as much as 1800 cpm per oocyte into each of the two Asn-linked oligosaccharides of this glycoprotein. Overall, about 50\% of the microinjected GDP-[3,4-3H]mannose was incorporated into Asn-linked oligosaccharides, a 10-fold greater value than that obtained when [2-3H]mannose was microinjected. Less than 10\% of the injected GDP-[3,4-3H]mannose was metabolized to water or converted to amino acids. This technique should facilitate studies of Asn-linked oligosaccharide biosynthesis, processing, and structure in recombinant proteins synthesized in Xenopus oocytes.",

author = "Cantor, \{Alan B.\} and Stuart Kornfeld",

note = "Funding Information: structural analysis, and Debbie Sinak for assistance with preparation of this manuscript. A.B.C. was supported by a National Institutes of Health Research Award GM-07200.Medical Scientist from the National Institutes of General Medical Sciences. This work was supported in part by United States Public Health Service Grant CA 08759.",

year = "1992",

month = sep,

doi = "10.1016/0003-2697(92)90427-9",

language = "English",

volume = "205",

pages = "220--226",

journal = "Analytical Biochemistry",

issn = "0003-2697",

number = "2",

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T1 - A method for [3H]mannose labeling of Asn-linked oligosaccharides on recombinant glycoproteins synthesized in Xenopus oocytes

AU - Cantor, Alan B.

AU - Kornfeld, Stuart

N1 - Funding Information: structural analysis, and Debbie Sinak for assistance with preparation of this manuscript. A.B.C. was supported by a National Institutes of Health Research Award GM-07200.Medical Scientist from the National Institutes of General Medical Sciences. This work was supported in part by United States Public Health Service Grant CA 08759.

PY - 1992/9

Y1 - 1992/9

N2 - We have developed an efficient method for labeling the Asn-linked oligosaccharides of recombinant glycoproteins synthesized in Xenopus laevis oocytes. By coinjecting GDP-[3,4-3H]mannose with mRNA for human cathepsin D, it was possible to incorporate as much as 1800 cpm per oocyte into each of the two Asn-linked oligosaccharides of this glycoprotein. Overall, about 50% of the microinjected GDP-[3,4-3H]mannose was incorporated into Asn-linked oligosaccharides, a 10-fold greater value than that obtained when [2-3H]mannose was microinjected. Less than 10% of the injected GDP-[3,4-3H]mannose was metabolized to water or converted to amino acids. This technique should facilitate studies of Asn-linked oligosaccharide biosynthesis, processing, and structure in recombinant proteins synthesized in Xenopus oocytes.

AB - We have developed an efficient method for labeling the Asn-linked oligosaccharides of recombinant glycoproteins synthesized in Xenopus laevis oocytes. By coinjecting GDP-[3,4-3H]mannose with mRNA for human cathepsin D, it was possible to incorporate as much as 1800 cpm per oocyte into each of the two Asn-linked oligosaccharides of this glycoprotein. Overall, about 50% of the microinjected GDP-[3,4-3H]mannose was incorporated into Asn-linked oligosaccharides, a 10-fold greater value than that obtained when [2-3H]mannose was microinjected. Less than 10% of the injected GDP-[3,4-3H]mannose was metabolized to water or converted to amino acids. This technique should facilitate studies of Asn-linked oligosaccharide biosynthesis, processing, and structure in recombinant proteins synthesized in Xenopus oocytes.

UR - https://www.scopus.com/pages/publications/0026616764

U2 - 10.1016/0003-2697(92)90427-9

DO - 10.1016/0003-2697(92)90427-9

M3 - Article

C2 - 1443567

AN - SCOPUS:0026616764

SN - 0003-2697

VL - 205

SP - 220

EP - 226

JO - Analytical Biochemistry

JF - Analytical Biochemistry

IS - 2

ER -

A method for [³H]mannose labeling of Asn-linked oligosaccharides on recombinant glycoproteins synthesized in Xenopus oocytes

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