tb-500 mastery course
Unit 2 of 12

molecular structure

the 43-amino-acid sequence, three functional domains, and what makes the LKKTET motif unique

43 Amino Acids, Three Domains

TB-500 is the 43-residue sequence of thymosin beta-4: an N-terminal signaling stretch that releases the Ac-SDKP anti-fibrotic fragment, the central LKKTET actin-binding motif that sequesters G-actin with a Kd of ~0.7 microM, and a C-terminal tail that enhances binding affinity and drives ILK-mediated migration signaling.

The label technically names a 7-residue LKKTET-Q heptapeptide, but vendors ship the full TB4 chain under it.

43 aa
full TB4 length
LKKTET
actin-binding motif
Ac-SDKP
N-terminal cleavage product
3
functional domains

meet the molecule

click any residue to see its role.

TB-500 / thymosin beta-4 domain map

Explore the Structure

tap a domain to see its role.

interactive structure viewer

molecular structure -- the simple version

what TB-500 is made of and why its shape matters, explained from scratch.

TB-500 is a chain of 43 amino acids (the building blocks that make up all proteins). It is a synthetic copy of a natural molecule called thymosin beta-4 that your body already produces in nearly every cell. The chain has three working sections: a front portion that releases a small anti-scarring fragment called Ac-SDKP (a four-amino-acid piece that reduces scar tissue formation); a middle section containing the LKKTET motif (a six-amino-acid sequence that grabs onto actin, the protein cells use as internal scaffolding); and a tail that wraps around actin to lock the grip in place. Unlike most proteins, TB-500 does not fold into a rigid shape -- it stays flexible, which lets it wrap around actin from multiple angles and interact with several different partner proteins.

A advanced: the LKKTET binding chemistry term
The LKKTET hexapeptide at residues 17-22 is the core functional element of TB-500. Leucine 17 provides a hydrophobic anchor inserting into a nonpolar pocket between actin subdomains 1 and 3. Lysines 18 and 19 form electrostatic salt bridges with Glu167 and Asp288 on actin. Threonines 20 and 22 make hydrogen bonds with subdomain 1 surfaces. The isolated LKKTET fragment retains actin-binding activity but with approximately 10-fold lower affinity than full-length Tb4. This motif is 100% conserved across all vertebrates spanning roughly 450 million years of evolution.
advanced: intrinsically disordered protein (IDP) properties
TB-500 belongs to the family of intrinsically disordered proteins (IDPs) -- molecules that function without a fixed three-dimensional fold. Its high charge density (multiple lysines, glutamic acids, aspartic acids), low hydrophobicity, and absence of cysteine residues (no disulfide bonds) prevent stable folding. The Zarbock et al. (1990) NMR study detected only a transient alpha-helical population of 10-20% at residues 5-16. This disorder enables binding promiscuity: Tb4 wraps around G-actin in one conformation, interacts with ILK/PINCH through another, and its Ac-SDKP fragment adopts yet another structure.
advanced: the three functional domains and their synergy
Domain 1 (residues 1-16) includes the Ac-SDKP tetrapeptide released by prolyl oligopeptidase cleavage, providing independent anti-fibrotic and anti-inflammatory signaling. Domain 2 (residues 17-30) contains the LKKTET actin-binding motif and is sufficient for G-actin sequestration alone. Domain 3 (residues 31-43) wraps along actin subdomain 2, enhancing binding affinity approximately 3-fold versus the central domain alone, and mediates ILK/PINCH interactions that drive cell migration via Akt. Fan et al. (2009) showed deleting domain 3 preserved actin binding but significantly reduced migration-promoting activity.

key terms

structural vocabulary you will need throughout this unit.

I intrinsically disordered protein structural concept
a protein that lacks a stable three-dimensional fold under physiological conditions. TB4 belongs to this class -- its high charge density, low hydrophobicity, and absence of disulfide bonds prevent folding. this disorder enables binding promiscuity: TB4 can wrap around actin in one conformation, contact ILK in another, and present Ac-SDKP in yet another.
D dissociation constant (Kd) binding affinity
a measure of how tightly two molecules bind. lower Kd = tighter binding. TB4 binds G-actin with a Kd of ~0.5-0.7 microM, meaning at the micromolar concentrations found inside cells (0.1-0.5 mM TB4), essentially all available G-actin is occupied by TB4.
A Ac-SDKP peptide fragment
N-acetyl-seryl-aspartyl-lysyl-proline -- the four-amino-acid N-terminal fragment cleaved from TB4 by prolyl oligopeptidase. Ac-SDKP has independent bioactivity: it reduces fibrosis, promotes angiogenesis, and is endogenously regulated by ACE. ACE inhibitors raise Ac-SDKP levels, possibly contributing to their cardioprotective effects.
S subdomain structural region
actin is divided into four subdomains (1-4). the LKKTET motif of TB4 contacts primarily subdomain 1, with additional contacts on subdomains 2 and 3. subdomain 1 contains Glu167 and Asp288, which form salt bridges with the lysine residues K18 and K19 of LKKTET.