Aspen SP1, an exceptional thermal, protease and detergent‐resistant self‐assembled nano‐particle

Abstract

Stable protein 1 (SP1) is a homo-oligomeric protein isolated from aspen (Populus tremula aspen) plants which forms a ring-shape dodecameric particle with a central cavity. The oligomeric form of SP1 is an exceptionally stable structure that is resistant to proteases (e.g., trypsin, V8, and proteinase K), high temperatures, organic solvents, and high levels of ionic detergent. Analytical ultra-centrifugation, chemical cross-linking, matrix-assisted laser-desorption time-of-flight mass spectrometry (MALDI-TOF-MS), and transmission electron microscopy were used to further characterize the SP1 dodecamer. Introduction of a single cysteine at the N-terminus of SP1 enabled the formation of disulfide bridges within the SP1 dodecamer, concurrent with increased melting point. A six-histidine tag was introduced at the N-terminus of SP1 to generate 6HSP1, and the DeltaNSP1 mutant was generated by a deletion of amino acids 2-6 at the N-terminus. Both 6HSP1 and DeltaNSP1 maintained their ability to assemble a stable dodecamer. Remarkably, these SP1 homo-dodecamers were able to re-assemble into stable hetero-dodecamers following co-electro-elution from SDS-PAGE. The exceptional stability of the SP1-nano ring and its ability to self-assemble hetero-complexes paves the way to further research in utilizing this unique protein in nano-biotechnology.

Enhanced cellulose degradation by nano-complexed enzymes: Synergism between a scaffold-linked exoglucanase and a free endoglucanase

“…The versatile nature of the cohesin-dockerin interaction on the cellulosomal scaffoldin provides a virtually unlimited number of arrays that can be used for fabrication of defined nanostructures for nanotechnological application (Ding et al, 2003). The unique cellulosomal cohesin-dockerin interaction was utilized to build an array of selfassembled quantum dots on a rigid cellulose crystal template (Ding et al, 2005) and the presentation of enzymatic modules on an alternative molecular scaffold: the stable protein 1 (SP1) from Populus tremula (Dgany et al, 2004;Heyman et al, 2007b;Wang et al, 2003Wang et al, , 2006Wang et al, , 2002.…”

Identifying water stress-response mechanisms in citrus by in silico transcriptome analysis

“…Nevertheless, SP1 is stress-related and has high thermostability as small heat shock proteins (sHSP), it does not present amino acid sequence nor function similarity in stress protection (Dgany et al, 2004). Therefore, Wang et al (2006) have described aspen SP1 as a remarkably resistant protein. It is boiling-stable and resistant to proteases, organic solvents and high levels of ionic detergent.…”

Protein Scaffold Engineering Towards Tunable Surface Attachment

“…SiSP1 was expressed in the bacteria-soluble fraction and formed a dodecamer complex with high stability characteristics (Supporting Information, Figure S2 a) and structural motifs as the wt SP1. [4][5][6][7] As mentioned above, wt SP1 and its mutants are stable in the presence of chaotropic agents, which denature most proteins. This unique feature was used to control the exposure of the inaccessible peptides in the SiSP1 mutant.…”

“…Overcoming the need for surface modifications combined with control of the protein surface affinity would enable the exploitation of protein immobilization for new materials and will increase future fabrication throughput. SP1, a ring-like protein that is highly stable to boiling and protease resistant, [4][5][6][7] was recently proposed as a new selfassembled molecular scaffold for nanobiotechnology and biomaterials applications. [8][9][10][11] Herein we present a novel strategy to control the interfacial adsorption of SP1 to an unmodified surface with high selectivity and controlled affinity.…”

“…The two mutants were expressed in E. coli and have high stability characteristics (Supporting Information, Figure S1) and complex formation as the wt SP1. [6,7] The position effect of the single-point mutation on the surface binding capability was analyzed by investigating the affinity of the two mutants to ultraflat gold surfaces (roughness less than 5 on an area of 5 mm 2 ). [12] Dynamic-mode atomic force microscopy (AFM) topographic imaging followed by flooding image analysis [13] ( Figure 1 d-f) was used to determine the percentage of the surface covered by the protein.…”