Archaeal Tetraether Liposomes Parkson L. Chong, Temple University, DMR The main goal of this research is to unravel the physical properties of liposomes made of archaeal bipolar tetraether lipids (BTL), so that we can improve the current usage of BTL liposomes and explore their new applications. We focused on liposomes composed of the polar lipid fraction E (PLFE), which is one of the main BTL isolated from the thermoacidophilic crenarchaeon Sulfolobus acidocaldarius. This organism thrives at o C and a pH of 2-3. PLFE is a mixture containing (a) GDGT (or caldarchaeol) and (b) GDNT (or calditolglycerocaldarchaeol) (illustrated right). These lipids have a pair of 40-carbon biphytanyl chains; each chain contains up to four cyclopentane rings. It is known that a decrease in archaeal cell growth temperature (T g ) decreases the number of cyclopentane rings in tetraether lipids. We have recently studied the effect of T g, inferentially the number of cyclopentane rings, on the physical properties of tetraether liposomes. Specifically, we have determined the compressibilities and volume fluctuations of PLFE liposomes derived from three different growth temperatures (T g ) by using calorimetry and molecular acoustics. The compressibility (not shown) and volume fluctuation (Figure 1) values of PLFE liposomes are low, compared to those found in a gel state of DPPC, and exhibit small but significant differences with T g. Among the three employed growth temperatures, the growth temperature 76 o C leads to the least compressible, and inferentially the most tightly packed PLFE lipid membranes. Note that 76 o C is in the temperature range for optimal growth of S. acidocaldarius (75-80 o C). This finding suggests that membrane packing in PLFE liposomes may actually vary with the number of cyclopentane rings in a non-linear manner, reaching maximal tightness when the lipids are derived from cells grown at the optimal growth temperaures. The main goal of this research is to unravel the physical properties of liposomes made of archaeal bipolar tetraether lipids (BTL), so that we can improve the current usage of BTL liposomes and explore their new applications. We focused on liposomes composed of the polar lipid fraction E (PLFE), which is one of the main BTL isolated from the thermoacidophilic crenarchaeon Sulfolobus acidocaldarius. This organism thrives at o C and a pH of 2-3. PLFE is a mixture containing (a) GDGT (or caldarchaeol) and (b) GDNT (or calditolglycerocaldarchaeol) (illustrated right). These lipids have a pair of 40-carbon biphytanyl chains; each chain contains up to four cyclopentane rings. It is known that a decrease in archaeal cell growth temperature (T g ) decreases the number of cyclopentane rings in tetraether lipids. We have recently studied the effect of T g, inferentially the number of cyclopentane rings, on the physical properties of tetraether liposomes. Specifically, we have determined the compressibilities and volume fluctuations of PLFE liposomes derived from three different growth temperatures (T g ) by using calorimetry and molecular acoustics. The compressibility (not shown) and volume fluctuation (Figure 1) values of PLFE liposomes are low, compared to those found in a gel state of DPPC, and exhibit small but significant differences with T g. Among the three employed growth temperatures, the growth temperature 76 o C leads to the least compressible, and inferentially the most tightly packed PLFE lipid membranes. Note that 76 o C is in the temperature range for optimal growth of S. acidocaldarius (75-80 o C). This finding suggests that membrane packing in PLFE liposomes may actually vary with the number of cyclopentane rings in a non-linear manner, reaching maximal tightness when the lipids are derived from cells grown at the optimal growth temperaures. Figure 1: Membrane volume fluctuations of PLFE liposomes derived from cells grown at 68°C (■), 76°C (○), and 81°C (∆), measured at various temperatures ranging from o C. Solid line: DPPC (dipalmitoyl-L-  -phosphatidylcholine, a diester lipid) liposomes for comparison.

Archaeal Tetraether Liposomes Parkson L. Chong, Temple University, DMR Broader Impacts on Science and Technology Archaeal bipolar tetraether lipids (BTL) are appealing biomaterials that hold great promise for technological applications. BTLs can be used as a stable lipid matrix for biosensors, a high temperature resistant lubricant, a light harvesting device, and nanoparticles for targeted imaging and therapy. Our study (as described in Slide 1) provides detailed and quantitative information about membrane packing in BTL liposomes, which will help improve the current knowledge and usage of BTL liposomes and explore their new applications. The new data (Figure 1) improve our understanding of the extraordinary stability of BLT liposomes against temperature. Our data showed that relative volume fluctuations of PLFE liposomes at any given temperature examined were much more damped than those found in DPPC liposomes. Volume fluctuations are closely related to solute permeation across lipid membranes and lateral motion of membrane components. Thus, the low values of relative volume fluctuations explain why PLFE liposomes exhibit unusually low proton permeation and dye leakage as well as limited lateral mobility, especially at low temperatures (e.g., < 26 o C) as shown in our previous publications. Our data showed that membrane packing in PLFE liposomes reaches maximal tightness when the tetraether lipids are derived from cells grown at the optimal growth temperaures. This information is important for future designs of extraordinarily stable BTL liposomes or lipid membranes for technological applications. Broader Impacts on Science and Technology Archaeal bipolar tetraether lipids (BTL) are appealing biomaterials that hold great promise for technological applications. BTLs can be used as a stable lipid matrix for biosensors, a high temperature resistant lubricant, a light harvesting device, and nanoparticles for targeted imaging and therapy. Our study (as described in Slide 1) provides detailed and quantitative information about membrane packing in BTL liposomes, which will help improve the current knowledge and usage of BTL liposomes and explore their new applications. The new data (Figure 1) improve our understanding of the extraordinary stability of BLT liposomes against temperature. Our data showed that relative volume fluctuations of PLFE liposomes at any given temperature examined were much more damped than those found in DPPC liposomes. Volume fluctuations are closely related to solute permeation across lipid membranes and lateral motion of membrane components. Thus, the low values of relative volume fluctuations explain why PLFE liposomes exhibit unusually low proton permeation and dye leakage as well as limited lateral mobility, especially at low temperatures (e.g., < 26 o C) as shown in our previous publications. Our data showed that membrane packing in PLFE liposomes reaches maximal tightness when the tetraether lipids are derived from cells grown at the optimal growth temperaures. This information is important for future designs of extraordinarily stable BTL liposomes or lipid membranes for technological applications. Other Broader Impacts International collaboration: ● This project was in collaboration with Professor Roland Winter’s group at University of Dortmund, Germany. Training undergraduate/graduate students: ● This project has been used to train several undergraduate students at Temple University in the academic year. ● One of the undergraduate students (Leeandrew Taylor) has made significant contributions to the work described in the previous slide and became a co-author in the resulting publication (Langmuir (2012) volume 28, pages ). ● One student (Nicole Haloupek) gave a poster presentation at the annual Biophysical Society meeting in San Diego, California, in ● This project has supported one summer (2012) intern through the collaboration with the Temple University Undergraduate Research program. ● Two PhD graduate students participated in this research project. Training students from under-represented groups: ● Three undergraduate students from the minority groups participated in this research project in the academic year.