Heme binding to human alpha-1 proteinase inhibitor

Heme is a unique prosthetic group of various hemoproteins that perform diverse biological functions; however, in its free form heme is intrinsically toxic in vivo. Due to its potential toxicity, heme binding to plasma proteins is an important safety issue in regard to protein therapeutics derived from human blood. While heme binding by hemopexin, albumin and α(1)-microglobulin has been extensively studied, the role of other plasma proteins remains largely unknown.

We examined two acute-phase plasma proteins, haptoglobin (Hp) and alpha-1 proteinase inhibitor (α(1)-PI) for possible interactions with heme and bilirubin (BR), the final product of heme degradation, using various techniques: UV/Vis spectroscopy, fluorescence, circular dichroism (CD), and surface plasmon resonance (SPR).

According to our data, Hp exhibits a very weak association with both heme and BR; α(1)-PI's affinity to BR is also very low. However, α(1)-PI's affinity to heme (K(D) 2.0×10(-8)M) is of the same order of magnitude as that of albumin (1.26×10(-8)M). The data for α(1)-PI binding with protoporphyrin IX (PPIX) suggest that the elimination of the iron atom from the porphyrin structure results in almost 350-fold lower affinity (K(D) 6.93×10(-6)M), thus indicating that iron is essential for the heme coordination with the α(1)-PI.

This work demonstrates for the first time that human α(1)-PI is a heme binding protein with an affinity to heme comparable to that of albumin.

Our data may have important implications for safety and efficacy of plasma protein therapeutics.