Chat with Fred and Chris Nov. 19 th, 2010 rhombohedra: SEM observation vs time, 24h-72h Bodega Marine Station payments - order Mytilus californianus spicule.

chat with Fred and ChrisNov. 19th, 2010

• rhombohedra: SEM observation vs time, 24h-72h

• Bodega Marine Station payments - order Mytilus californianus

• spicule matrix proteins: produce in yeast (Liam Holt), then harvest and isolate with TAP (twice ).

• which proteins do we try?• can we go for the 5 most abundant in spicules?

• do we agree about the “protein-function” assay on calcite polished crystals? It only shows ACC-stabilization function.

• other, better ideas?

Mechanism of transformation of amorphous calcium carbonate into calcite in the sea urchin larval spiculeY Politi, RA Metzler, M Abrecht, B Gilbert, F Wilt, I Sagi, L Addadi, S Weiner, and PUPA GilbertProcs. Natl. Acad. Sci. USA 105, 17362-17366 (2008)

sea urchin spicules revealed 3 phases : 2 Amorphous Calcium Carbonate (ACC), and 1 biogenic calcite: type 1, type 2, type 3.

type 1type 2type 3?

spicule

shadow

Si wafer

X-PEEM (photoelectron emission microscopy) image and spectra

Transformation and crystallization energetics of synthetic and biogenic ACCAV Radha, TZ Forbes, CE Killian, PUPA Gilbert, and A NavrotskyProcs. Natl. Acad. Sci. USA, 107, 16438-16443, 2010.

transitions type 12 and type 23 are both energetically downhill

isothermal acid-solution calorimetry data

red = hydrated ACCgreen = anhydrous ACCblue = crystalline CaCO3

transitions type 12 and type 23 are both energetically downhill

isothermal acid-solution calorimetry data

red = hydrated ACCgreen = anhydrous ACCblue = crystalline CaCO3

Transformation and crystallization energetics of synthetic and biogenic ACCAV Radha, TZ Forbes, CE Killian, PUPA Gilbert, and A NavrotskyProcs. Natl. Acad. Sci. USA, 107, 16438-16443, 2010.

transitions type 12 and type 23 are both energetically downhill

isothermal acid-solution calorimetry data

red = hydrated ACCgreen = anhydrous ACCblue = crystalline CaCO3

Transformation and crystallization energetics of synthetic and biogenic ACCAV Radha, TZ Forbes, CE Killian, PUPA Gilbert, and A NavrotskyProcs. Natl. Acad. Sci. USA, 107, 16438-16443, 2010.

red = type 1, hydrated ACCgreen = type 2, anhydrous ACCblue = type 3, biogenic calcite

R+G=YellowR+B=MagentaG+B=Cyan

Component Analysis of X-PEEM data (X-ray PhotoElectron Emission spectroMicroscopy)

CaCO3 phase transitions in the sea urchin larval spiculeYUT Gong, CE Killian, IC Olson, NP Appathurai, RA Metzler, AL Amasino, FH Wilt, and PUPA Gilbertin preparation

type 1type 2type 3 sequence experimentally confirmed in spicule cross-section (20-nm pixels)

500 nm

type 12 and 23 are fundamentally different transitions

red = type 1, hydrated ACCgreen = type 2, anhydrous ACCblue = type 3, biogenic calcite

R+G+B=WhiteR+G=YellowG+B=CyanR+B=Magenta

spicule cross-section (20-nm pixels)Component Analysis of X-PEEM data(X-ray PhotoElectron Emission spectroMicroscopy)

CaCO3 phase transitions in the sea urchin larval spiculeYUT Gong, CE Killian, IC Olson, NP Appathurai, RA Metzler, AL Amasino, FH Wilt, and PUPA Gilbertin preparation

RGB enabled

red = type 1, hydrated ACCgreen = type 2, anhydrous ACCblue = type 3, biogenic calcite

simulation modelsRG inhibited

type 12 and 23 are fundamentally different transitions

CaCO3 phase transitions in the sea urchin larval spiculeYUT Gong, CE Killian, IC Olson, NP Appathurai, RA Metzler, AL Amasino, FH Wilt, and PUPA Gilbertin preparation

RGB enabled

1 µm

red = type 1, hydrated ACCgreen = type 2, anhydrous ACCblue = type 3, biogenic calcite

experimental data simulation models

RG inhibited

type 12 and 23 are fundamentally different transitions

type 12 transition is biologically controlledtype 23 transition is spontaneous

CaCO3 phase transitions in the sea urchin larval spiculeYUT Gong, CE Killian, IC Olson, NP Appathurai, RA Metzler, AL Amasino, FH Wilt, and PUPA Gilbertin preparation

RGB map

Green edge

Red edge

Magenta nanoparticles

Hydrated ACCAnhydrous ACCBiogenic calcite

red or green edges

magenta nanoparticles

Yutan: I see just as many cyan nanoparticles as magenta nanoparticles.What did you change here?are the above slides no longer valid?

48h and 72h spicules were extracted and deminineralized with acetic acid,neutralized with Tris buffer at pH 8, then dialyzed against DDH2O

sanity check:a droplet of DDH2O was deposited on a calcite crystal, then air-dried

sanity check:a droplet of DDH2O was deposited on a calcite crystal, then air-dried

re-crystallized calcite is more disordered:lower peaks 2 and 4