Biological activities of histidine-rich peptides; merging biotechnology and nanomedicine

Abstract

Histidine-rich peptides are commonly used in recombinant protein production as purification tags, allowing the one-step affinity separation of the His-tagged proteins from the extracellular media or cell extracts. Genetic engineering makes feasible the post-purification His-tag removal by inserting, between the tag and the main protein body, a target site for trans-acting proteases or a self-proteolytic peptide with regulatable activities. However, for technical ease, His tags are often not removed and the fusion proteins eventually used in this form. In this commentary, we revise the powerful biological properties of histidine-rich peptides as endosomolytic agents and as architectonic tags in nanoparticle formation, for which they are exploited in drug delivery and other nanomedical applications. These activities, generally unknown to biotechnologists, can unwillingly modulate the functionality and biotechnological performance of recombinant proteins in which they remain trivially attached.

Sheltering DNA in self-organizing, protein-only nano-shells as artificial viruses for gene delivery

“…19 When nanoparticles formed by R9-GFP-H6 at pH 7 and 8 ( Figure 1, A) were incubated with DNA, particle size remained close to 20 nm (Figure 1, B), the size previously observed in the absence of DNA. 15 At pH 4 and 10, protein-DNA complexes peaked at 0.8 and 2 μm respectively (Figure 1, B), which is in agreement with the tendency of the protein alone to form amorphous aggregates under denaturing conditions (Figure 1, A).…”

“…H6 (HHHHHH) is at the same time a useful tag for one-step chromatographic protein purification and a potent endosomolytic agent. 19 Precise amino acid sequences at the links between GFP and the fused peptides can be found elsewhere. 17 The protein constructs indicated above were produced in bacteria following conventional procedures and purified in a single step by His-based affinity chromatography, 15 through activities assisted by the Protein Production Platform (CIBER-BBN) (http://www.bbn.ciber-bbn.es/programas/plataformas/ equipamiento).…”

Intrinsic functional and architectonic heterogeneity of tumor-targeted protein nanoparticles

“…In receptor-targeted nanoparticles that are activated with overhanging peptides, uptake might be in addition modulated by the exposure and bioavailability of functional ligands on the particle's surface. In our system, these ligands are the tumor homing peptides A5G27 and T22, while the H6 tail has an important role in the endosomal escape upon internalization [35]. The variability in the specific GFP fluorescence emission when comparing all the nanoparticles studied here (Figure 1 B) suggested alternative configurations of the material [22].…”

Intracellular CXCR4+ cell targeting with T22-empowered protein-only nanoparticles

“…The efficient endosomal escape of the constructs generated might be due to the proton-sponge activity of the accompanying polyhistidines that, while useful for one-step protein purification, act also as a proton sponge, permitting endosomal disruption and delivery of the functionalized materials into the cytoplasm. 22 On the other hand, when tested in an in vivo animal model of colorectal cancer, in which CXCR4…”

“…However, the relative low proportion of yellow signals and rapid accumulation of nanoparticles close to the nuclear region through fast cytoplasmic trafficking ( Figure 4D) were indicative of early endosomal escape. This was probably promoted by the accompanying hexahistidine tag, that has powerful endosomolytic properties, 22 inducing early endosomal escape in related protein-only nanoparticles.…”