Expression in Pichia pastoris and characterization by circular dichroism and NMR of rhodostomin

Proteins
R T GuoW J Chaung

Abstract

Rhodostomin (Rho) is a snake venom protein isolated from Calloselasma rhodostoma. Rho is a disintegrin that inhibits platelet aggregation by blocking the binding of fibrinogen to the integrin alpha(IIb)beta3 of platelets. Rho produced in Escherichia coli inhibited platelet aggregation with a K(I) value of 263 nM. Although functional, Rho produced in E. coli is misfolded based on our 2D and 3D NMR studies. In order to correct the folding problem, Rho was expressed in Pichia pastoris. The recombinant Rho expressed in P. pastoris inhibited platelet aggregation with a resulting K(I) value of 70 nM. This is the same potency as that of native Rho. CD analysis showed that the secondary structures of Rho are pH-independent and contain 3.5-7.9% alpha-helix, 48.2-50.5% beta-structures, and 42.3-47% coil. The sequential assignment and structure analysis of Rho were obtained using 2D and 3D 15N-edited NMR spectra. These results provide the first direct evidence that highly disulfide-bonded disintegrin can be expressed in P. pastoris with the correct fold. This evidence may serve as the basis for exploring the structure and function relationships as well as the dynamics of disintegrin and its variants.

References

Oct 1, 1992·Current Opinion in Cell Biology·C P Blobel, J M White
Nov 15, 1994·Structure·C E WhiteE A Komives
Oct 28, 1998·Proceedings of the Society for Experimental Biology and Medicine·M A McLaneS Niewiarowski
Jan 29, 1999·Lancet·E J TopolE F Plow

❮ Previous
Next ❯

Citations

Feb 24, 2010·Amino Acids·Irina VetterGlenn F King
Apr 9, 2008·Acta Crystallographica. Section D, Biological Crystallography·Natalia MoiseevaMarc Allaire
Mar 23, 2002·Protein Science : a Publication of the Protein Society·Vladimir N Uversky
Sep 4, 2010·Toxicon : Official Journal of the International Society on Toxinology·Raveendra AnangiWoei-Jer Chuang
Sep 18, 2007·Toxicon : Official Journal of the International Society on Toxinology·Pon Singhamatr, Ponlapat Rojnuckarin
Oct 4, 2012·Protein Science : a Publication of the Protein Society·Chun-Ho ChengWoei-Jer Chuang
Jan 26, 2006·Comparative Biochemistry and Physiology. Toxicology & Pharmacology : CBP·O H P Ramos, H S Selistre-de-Araujo
Oct 4, 2005·Biochemical Pharmacology·Yu-Ting LinWen-Mei Fu
Sep 18, 2012·Toxicon : Official Journal of the International Society on Toxinology·Yi-Chun ChenWoei-Jer Chuang
Jan 2, 2014·Toxicon : Official Journal of the International Society on Toxinology·Panchalee Jangprasert, Ponlapat Rojnuckarin
Aug 9, 2018·Toxins·Victor DavidRussolina Benedeta Zingali
Mar 14, 2013·The Journal of Clinical Endocrinology and Metabolism·Hsiu-Mei ChenShaw-Jenq Tsai
Aug 18, 2017·Journal of Thrombosis and Haemostasis : JTH·Y-J KuoT-F Huang

❮ Previous
Next ❯

Related Concepts

Related Feeds

Adhesion Molecules in Health and Disease

Cell adhesion molecules are a subset of cell adhesion proteins located on the cell surface involved in binding with other cells or with the extracellular matrix in the process called cell adhesion. In essence, cell adhesion molecules help cells stick to each other and to their surroundings. Cell adhesion is a crucial component in maintaining tissue structure and function. Discover the latest research on adhesion molecule and their role in health and disease here.

Cajal Bodies & Gems

Cajal bodies or coiled bodies are dense foci of coilin protein. Gemini of Cajal bodies, or gems, are microscopically similar to Cajal bodies. It is believed that Cajal bodies play important roles in RNA processing while gems assist the Cajal bodies. Find the latest research on Cajal bodies and gems here.