Transforming Growth Factor-β1 (TGF-β1) Characterization

Introduction to Transforming Growth Factor-β1 (TGF-β1)

The transforming growth factor-beta (TGF-β) family of proteins has emerged as a particularly noteworthy group due to its wide-ranging multifunctional capabilities. Notably, one member of this family, namely transforming growth factor-beta1 (TGF-β1), was initially identified for its ability to induce a phenotypic transformation in rat fibroblasts.

Fig 1. Architecture of proTGF-β1. (Shi, Minlong, et al. 2011)

Since its discovery, TGF-β1 has been found to exert significant regulatory actions on both normal and neoplastic cells. Importantly, nearly all cell types possess specific high-affinity receptors for TGF-β, and multiple cell types are capable of synthesizing this protein. Consequently, TGF-β stands as a pivotal regulatory molecule that can elicit its effects through autocrine and paracrine mechanisms.

Structure

Mature TGF-β1 is composed of two identical peptide chains, each consisting of 112 amino acids. The protein is a homodimer with a molecular weight of 24 kDa and contains a total of nine disulfide bonds. One of these bonds links the monomeric subunits together. When reduced, the homodimer dissociates into two identical peptides, each weighing approximately 12 kDa.

The nonreduced form of the homodimer is the biologically active form of the molecule. Although the crystal structure of TGF-β1 is currently unknown, it has been determined for TGF-β2. TGF-β1 shares 71% homology with TGF-β2 in terms of amino acid sequence, indicating a probable structural similarity between the two proteins.

Fig 2. A Sequence comparisons and 3-D-1-D profile scores for the TGF-β superfamily. (Daopin, S., et al. 1992)

Studies on TGF-β2 have shown that it lacks a well-defined hydrophobic core and instead exhibits an unusual elongated nonglobular fold. This fold has dimensions of approximately 60 Å by 20 Å by 15 Å. Sequence analysis of other members of the TGF-β superfamily suggests that they also adopt this unique fold.

Within the TGF-β1 protein, eight cysteines form four intrachain disulfide bonds, which are localized in a central core region. The dimerization of the protein is further stabilized by the ninth cysteine, which forms an interchain disulfide bond. Additionally, there are two identical hydrophobic interfaces that contribute to the stability of the dimeric structure.

Integrity of TGF-β1

RP-HPLC (Reverse Phase High Performance Liquid Chromatography) methodology is utilized to scrutinize the integrity of TGF-β1 by distinguishing TGF-β1 and its derivatives, grounded on the discrepancy in their partitioning among the hydrophobic matrix within the column and the comparatively hydrophilic mobile phase.

Fig 3. Reversed-Phase High-Performance Liquid Chromatography (RP-HPLC) of TGF-β1. (Pearlman, Rodney, and Y. John Wang, eds. 1996)

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References

1. Shi, Minlong, et al. Latent TGF-β structure and activation. Nature. 2011, 474.7351: 343-349.
2. Daopin, S., et al. Crystal structure of transforming growth factor-beta 2: an unusual fold for the superfamily. Science. 1992, 257(5068), 369–373.
3. Pearlman, Rodney, and Y. John Wang, eds. Formulation, characterization, and stability of protein drugs. Vol. 9. Springer Science & Business Media. 1996.

For research use only, not intended for any clinical use.