Engineering of Ribosome-inactivating Proteins for Improving Pharmacological Properties

Abstract

Ribosome-inactivating proteins (RIPs) are N-glycosidases, which depurinate a specific adenine residue in the conserved α-sarcin/ricin loop (α-SRL) of rRNA. This loop is important for anchoring elongation factor (EF-G for prokaryote or eEF2 for eukaryote) in mRNA translocation. Translation is inhibited after the attack. RIPs therefore may have been applied for anti-cancer, and anti-virus and other therapeutic applications. The main obstacles of treatment with RIPs include short plasma half-life, non-selective cytotoxicity and antigenicity. This review focuses on the strategies used to improve the pharmacological properties of RIPs on human immunodeficiency virus (HIV) and cancers. Coupling with polyethylene glycol (PEG) increases plasma time and reduces antigenicity. RIPs conjugated with antibodies to form immunotoxins increase the selective toxicity to target cells. The prospects for future development on the engineering of RIPs for improving their pharmacological properties are also discussed.

Keywords: anti-HIV; anti-cancer; immunotoxin; ribosome inactivating protein; therapeutic applications.

Figure 1

The cell cycle of HIV and the anti-HIV mechanism of representative RIPs. RIPs including TCS, GAP31, MAP30, PAP, marmorin and saporin can attack different steps of the life cycle of HIV and inhibit its growth. The mechanisms are not just due to rRNA depurination. TCS: trichosanthin; MAP30: momordica anti-HIV protein. GAP31: gelonium anti-HIV protein; PAP: pokeweed antiviral protein; MOD: maize ribosome-inactivating protein; DAP30: dianthus anti-HIV protein; MAPK: mitogen-activated protein kinase; HIV: human immunodeficiency virus.

Figure 2

Schematic diagram of the RTA and maize RIP variants. The recombinant maize RIP precursor pro-RIP contains a 25 amino acids internal inactivation region. RTA: ricin A chain; HIV: human immunodeficiency virus; TAT: transcriptional activator Tat protein. TAT-Pro-HIV-P2/NC and TAT-Pro-HIV-MA/CA: N-termini of pro-RIP were fused with a TAT sequence. First and last 10 aa in internal inactivation region were replaced by the HIV-1 recognition p2/NC site (TATIM/MQRGN) and the MA/CA site (VSQNY/PIVQN), respectively. RTA-C10: MA/CA site fused to C-terminal of RTA. RTA-C25: two MA/CA sites were fused and separated by MQMPE (middle five residues of pro-RIP).

Figure 3

Ricin-induced cell death in the anti-tumor process. Arrows represent the activation of receptors and blunt arrows represent inhibition of receptors. Pathways involved are stated in blue boxes. The inhibition of protein synthesis, cell apoptosis and the release of cytokine inflammatory mediators are considered as the possible mechanism. p38: p38 mitogen-activated protein kinases; JNK: c-Jun N-terminal kinase; MAPK: mitogen-activated protein kinase; PI3K: phosphoinositide 3-kinases; JAK2: Janus kinase 2; STAT: signal transducer and activator of transcription; PARP: poly ADP-ribose polymerase; iNOS: inducible nitric oxide synthase; TNF-α: tumor necrosis factor-α; IL: Interleukin; Bcl-xl: B-cell lymphoma-extra large; Bax: Bcl-2-associated X protein.