SCAFFOLD PROTEIN ASSEMBLY SERIES
SCAFFOLD PROTEIN ASSEMBLY FREE

Our team is researching naturally occurring scaffolds possessing the ability to spatially organize enzymes. Among these synthetic scaffolds, protein arrays and DNA nanostructures are the most biocompatible and have the potential to form the basis of a powerful platform to enhance multi-enzyme catalysis for biotechnology applications ( Klein et al., 2019 Lim et al., 2019). Artificial multi-enzyme scaffolds have been utilized both in vivo and in vitro to improve product production ( Siu et al., 2015 Ellis et al., 2019). Based on these concepts, various synthetic scaffolds aimed to spatially organize enzymes were designed and used for immobilizing enzymes to enhance their activity ( Conrado et al., 2008 Dueber et al., 2009 Fu et al., 2014).

SCAFFOLD PROTEIN ASSEMBLY SERIES

As a result, there is a preference for a folded state over an unfolded state and their close proximity allows them to execute a series of biocatalytic events more efficiently through substrate channeling ( Miles et al., 1999 Zhang, 2011).

SCAFFOLD PROTEIN ASSEMBLY FREE

Specifically, high concentration of enzymes in a confined environment are more stable than those free in solution. Our results demonstrate that the T4 phage capsid can act as a naturally occurring scaffold with substantial potential to enhance enzyme activity by spatially organizing enzymes on the capsid Hoc.Ĭatalytic properties of enzymes are greatly affected by their surrounding microenvironment, particularly enzymes retained in a small area either by limited surface or restricted volume ( Kuchler et al., 2016). Kinetic analysis also revealed that the immobilized three-enzyme cascade has an 18-fold higher converted number of NAD + to NADH relative to the mixtures in solution. The capsid-immobilized Maltase has a fourfold higher initial rate relative to Maltase free in solution. Covalent constructs between each of the enzymes and the outer capsid protein Hoc were prepared through SpyTag–Sp圜atcher pairing and assembled onto phage capsids in vitro with an estimated average of 90 copies per capsid. Herein, we examined the utility of the T4 phage capsid to serve as a naturally occurring protein scaffold for the immobilization of a three-enzyme cascade: Amylase, Maltase, and Glucokinase. The most bio-compatible synthetic scaffolds known for enzyme immobilization are protein and DNA nanostructures. Over the past two decades, various scaffolds have been designed and synthesized to organize enzyme cascades spatially for enhanced enzyme activity based on the concepts of substrate channeling and enhanced stability.

Center for Bio/Molecular Science and Engineering, Naval Research Laboratory, Washington, DC, United States.

The recent advance of DNA-binding adaptor mediated assembly of proteins on the DNA scaffolds is highlighted and discussed in connection with the future perspectives of protein assembled DNA nanoarchitectures.Jinny L. The assembling methods were categorized into two main classes, noncovalent and covalent conjugation, with both showing pros and cons. Here, we provide an overview of the existing methods applied for assembling proteins of interest on DNA scaffolds. The methods to arrange proteins of interest on DNA scaffolds at high yields while retaining their activity are still the most demanding task in constructing usable protein-modified DNA nanostructures. When loading proteins of interest on DNA nanostructures, the nanostructures by themselves act as scaffolds to specifically control the location and number of protein molecules. DNA nanostructures loaded with various types of proteins hold promise for applications in the life and material sciences. Among the molecules to functionalize DNA nanostructures, proteins are one of the most attractive candidates due to their vast functional variations. In addition to their structure and function, modification the DNA nanostructures by other molecules opens almost unlimited possibilities for producing functional DNA-based architectures. DNA is an attractive molecular building block to construct nanoscale structures for a variety of applications.

Scaffold protein assembly