Author: Saitoh, Koichi ; Nawara, Hend M. ; Alam, Md Jahangir ; Salomon, David S. ; H Zahra, Maram ; Afify, Said M. ; Seno, Akimasa ; Seno, Masaharu ; Sheta, Mona ; Hassan, Ghmkin ; Mansour, Hager ; Du, Juan ; Oh, Sue Young ; Xu, Yanning ; Abu Quora, Hagar A. ; Monzur, Sadia

AbstractHuman Cripto‐1 is a member of the epidermal growth factor (EGF)‐Cripto‐FRL‐1‐Cryptic (CFC) family family and performs critical roles in cancer and various pathological and developmental processes. Recently we demonstrated that a soluble form of Cripto‐1 suppresses the self‐renewal and enhances the differentiation of cancer stem cells (CSCs). A functional form of soluble Cripto‐1 was found to be difficult to obtain because of the 12 cysteine residues in the protein which impairs the folding process. Here, we optimized the protocol for a T7 expression system, purification from inclusion bodies under denatured conditions refolding of a His‐tagged Cripto‐1 protein. A concentrations of 0.2−0.4 mM isopropyl β‐D‐1‐thiogalactopyranoside (IPTG) at 37°C was found to be the optimal concentration for Cripto‐1 expression while imidazole at 0.5 M was the optimum concentration to elute the Cripto‐1 protein from a Ni‐column in the smallest volume. Cation exchange column chromatography of the Cripto‐1 protein in the presence of 8 M urea exhibited sufficient elution profile at pH 5, which was more efficient at recovery. The recovery of the protein reached to more than 26.6% after refolding with arginine. The purified Cripto‐1 exhibited high affinity to the anti‐ALK‐4 antibody and suppressed sphere forming ability of CSCs at high dose and induced cell differentiation.

01 Jul 2001·Biomedicine & PharmacotherapyQ2 · MEDICINE

Additive effect of tunicamycin and dextran sulfate on the binding of monoclonal antibody to the V2 domain of the envelope glycoprotein 120 of human immunodeficiency virus type 1

Q2 · MEDICINE

Article

Author: P.P Jagodzinski ; W.H Trzeciak

Using flow cytometry or immunoprecipitation analysis in cells chronically infected with HIV-1 IIIB Supt-1, we noticed an additive effect of tunicamycin and low molecular weight dextran (LMDS) on the binding of the G3-4 monoclonal antibody to monomeric and oligomeric forms of glycoprotein 120 (gp120). The inhibition of glycosylation by tunicamycin reduced the number of monomeric and oligomeric forms of gp120. The inhibition of the binding of the G3-4 antibody to monomeric and oligomeric forms of gp120 was more pronounced in the presence of LMDS. We also found that the G3-4 antibody can not recognise the nascent polypeptide chain of the envelope glycoprotein.

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Neoplasms	Discovery	NL	Synthon BV	-

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