Inefficient processing of human protein C in the mouse mammary gland

William N. Drohan, Da Wei Zhang, Rekhak Paleyanda, Rouling Chang, Marie Wroble, William Velander, Henryk Lubon

Research output: Contribution to journalArticle

38 Citations (Scopus)

Abstract

Vitamin K-dependent plasma protein, human Protein C (HPC) has been expressed in transgenic mice, using a 4.2kb mouse whey acidic protein (WAP) promoter, 9.0 kb HPC gene and 0.4 kb 3′flanking sequences. Expression was mammary gland-specific and the recombinant human Protein C (rHPC) was detected in milk at concentrations of 0.1 to 0.7mg ml-1. SDS-PAGE revealed that the single, heavy and light chains of rHPC migrated with increased electrophoretic mobility, as compared to HPC. Enzymatic deglycosylation showed that these molecular weight disparities are in part due to differential glycosylation. The substantial increase observed in the amount of single chain protein, as well as the presence of the propeptide attached to 20-30% of rHPC, suggest that mouse mammary epithelial cells are not capable of efficient proteolytic processing of rHPC. The Km of purified rHPC for the S-2366 synthetic substrate was similar to that of plasma-derived HPC, while the specific activity was about 42-77%. Amino acid sequence analyses and low anticoagulant activity of purified rHPC suggest that γ-carboxylation of rHPC is insufficient. These results show that proteolytic processing and γ-carboxylation can be limiting events in the overexpression of fully biologically active rHPC in the mouse mammary gland.

Original languageEnglish (US)
Pages (from-to)355-364
Number of pages10
JournalTransgenic Research
Volume3
Issue number6
DOIs
StatePublished - Nov 1 1994

Fingerprint

Human Mammary Glands
Protein C
mammary glands
Recombinant Proteins
mice
proteins
carboxylation
anticoagulant activity
Vitamin K
Protein S
Protein Sequence Analysis
vitamin K
Glycosylation
Anticoagulants
Transgenic Mice
glycosylation
Blood Proteins
Polyacrylamide Gel Electrophoresis
whey
Milk

Keywords

  • human protein C
  • mouse mammary gland
  • processing

ASJC Scopus subject areas

  • Biotechnology
  • Animal Science and Zoology
  • Agronomy and Crop Science
  • Genetics

Cite this

Drohan, W. N., Zhang, D. W., Paleyanda, R., Chang, R., Wroble, M., Velander, W., & Lubon, H. (1994). Inefficient processing of human protein C in the mouse mammary gland. Transgenic Research, 3(6), 355-364. https://doi.org/10.1007/BF01976767

Inefficient processing of human protein C in the mouse mammary gland. / Drohan, William N.; Zhang, Da Wei; Paleyanda, Rekhak; Chang, Rouling; Wroble, Marie; Velander, William; Lubon, Henryk.

In: Transgenic Research, Vol. 3, No. 6, 01.11.1994, p. 355-364.

Research output: Contribution to journalArticle

Drohan, WN, Zhang, DW, Paleyanda, R, Chang, R, Wroble, M, Velander, W & Lubon, H 1994, 'Inefficient processing of human protein C in the mouse mammary gland', Transgenic Research, vol. 3, no. 6, pp. 355-364. https://doi.org/10.1007/BF01976767
Drohan, William N. ; Zhang, Da Wei ; Paleyanda, Rekhak ; Chang, Rouling ; Wroble, Marie ; Velander, William ; Lubon, Henryk. / Inefficient processing of human protein C in the mouse mammary gland. In: Transgenic Research. 1994 ; Vol. 3, No. 6. pp. 355-364.
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