I. Astner1, J.O. Schulze1, W.-D. Schubert1, D. Jahn2, D.W. Heinz1
1 Division of Structural Biology, German Research Centre for Biotechnology, Mascheroder Weg 1, D-38124 Braunschweig, 2 Institute of Microbiology, Technical University Braunschweig, Spielmannstr. 7, D-38106 Braunschweig
The first dedicated enzyme in heme biosynthesis in humans is 5-aminolevulinate synthase (ALAS). eALAS (e for erythroid) is one of two isoforms of ALAS expressed in mammals and is responsible for approximately 90% of body heme production.
Naturally occurring mutations in human eALAS directly cause a class of porphyrias known as X-linked sideroblastic anaemia (XLSA). These disorders are characterized by inadequate formation of heme and accumulation of iron in erythroblast mitochondria.
Both human and bacterial (Rhodobacter capsulatus) ALAS were cloned and overexpressed in E. coli and purified to homogeneity. The bacterial ALAS, 50% identical by sequence to its human counterpart, crystallized successfully. The crystal structure was solved and refined to a resolution of 2.1 Ã….
The high resolution of the crystal structure of ALAS allows us to locate most naturally occurring mutations with high precision. Hence we can determine why these mutations affect the enzyme efficiency in producing aminolevulinic acid (ALA), a prerequisite intermediate in synthesizing heme.
Several mutations occur in the binding pockets of the two substrates (glycine and succinyl Co-A) and of the cofactor PLP (pyridoxal-5'-phosphate, product of Vitamin B6). Others are located in the channels leading from the protein surface to the active site, while yet others are located at the interface of the two subunits making up the functional ALAS dimer.
We are therefore now in a position to interpret the clinical XLSA-cases in terms of the three dimensional structure of the enzyme involved. Thereby new impetus is given to finding ways of treating XLSA other than by increasing levels of Vitamin B6 or the product aminolevulinc acid.