PMID- 27139003
OWN - NLM
STAT- MEDLINE
DCOM- 20170609
LR  - 20171218
IS  - 1520-4812 (Electronic)
IS  - 1043-1802 (Linking)
VI  - 27
IP  - 5
DP  - 2016 May 18
TI  - Dynamic Stabilization of Expressed Proteins in Engineered Diatom Biosilica 
      Matrices.
PG  - 1205-9
LID - 10.1021/acs.bioconjchem.6b00165 [doi]
AB  - Self-assembly of recombinant proteins within the biosilica of living diatoms 
      represents a means to construct functional materials in a reproducible and 
      scalable manner that will enable applications that harness the inherent 
      specificities of proteins to sense and respond to environmental cues. Here we 
      describe the use of a silaffin-derived lysine-rich 39-amino-acid targeting 
      sequence (Sil3T8) that directs a single chain fragment variable (scFv) antibody 
      or an enhanced green fluorescent protein (EGFP) to assemble within the biosilica 
      frustule, resulting in abundance of >200 000 proteins per frustule. Using either 
      a fluorescent ligand bound to the scFv or the intrinsic fluorescence of EGFP, we 
      monitored protein conformational dynamics, accessibility to external quenchers, 
      binding affinity, and conformational stability. Like proteins in solution, 
      proteins within isolated frustules undergo isotropic rotational motion, but with 
      2-fold increases in rotational correlation times that are indicative of weak 
      macromolecular associations within the biosilica. Solvent accessibilities and 
      high-affinity (pM) binding are comparable to those in solution. In contrast to 
      solution conditions, scFv antibodies within the biosilica matrix retain their 
      binding affinity in the presence of chaotropic agents (i.e., 8 M urea). Together, 
      these results argue that dramatic increases in protein conformational stability 
      within the biosilica matrices arise through molecular crowding, acting to retain 
      native protein folds and associated functionality with the potential to allow the 
      utility of engineered proteins under a range of harsh environmental conditions 
      associated with environmental sensing and industrial catalytic transformations.
FAU - Xiong, Yijia
AU  - Xiong Y
AD  - Department of Basic Medical Sciences, Western University of Health Sciences , 
      Lebanon, Oregon 97355, United States.
FAU - Ford, Nicole R
AU  - Ford NR
AD  - Marine Biotechnology Group, Pacific Northwest National Laboratory , Sequim, 
      Washington 98382, United States.
FAU - Hecht, Karen A
AU  - Hecht KA
AD  - Marine Biotechnology Group, Pacific Northwest National Laboratory , Sequim, 
      Washington 98382, United States.
FAU - Roesijadi, Guritno
AU  - Roesijadi G
AD  - Marine Biotechnology Group, Pacific Northwest National Laboratory , Sequim, 
      Washington 98382, United States.
AD  - Department of Microbiology, Oregon State University , Corvallis, Oregon 97331, 
      United States.
FAU - Squier, Thomas C
AU  - Squier TC
AD  - Department of Basic Medical Sciences, Western University of Health Sciences , 
      Lebanon, Oregon 97355, United States.
LA  - eng
PT  - Journal Article
PT  - Research Support, U.S. Gov't, Non-P.H.S.
DEP - 20160505
PL  - United States
TA  - Bioconjug Chem
JT  - Bioconjugate chemistry
JID - 9010319
RN  - 0 (Single-Chain Antibodies)
RN  - 0 (enhanced green fluorescent protein)
RN  - 147336-22-9 (Green Fluorescent Proteins)
RN  - 7631-86-9 (Silicon Dioxide)
SB  - IM
MH  - *Diatoms
MH  - *Engineering
MH  - Gene Expression
MH  - Green Fluorescent Proteins/*chemistry/*genetics
MH  - Models, Molecular
MH  - Protein Stability
MH  - Protein Structure, Secondary
MH  - Silicon Dioxide/*chemistry
MH  - Single-Chain Antibodies/*chemistry
EDAT- 2016/05/04 06:00
MHDA- 2017/06/10 06:00
CRDT- 2016/05/04 06:00
PHST- 2016/05/04 06:00 [entrez]
PHST- 2016/05/04 06:00 [pubmed]
PHST- 2017/06/10 06:00 [medline]
AID - 10.1021/acs.bioconjchem.6b00165 [doi]
PST - ppublish
SO  - Bioconjug Chem. 2016 May 18;27(5):1205-9. doi: 10.1021/acs.bioconjchem.6b00165. 
      Epub 2016 May 5.