MHC class III is a group of proteins belonging to the class of major histocompatibility complex. Unlike other MHC types such as MHC class I and MHC class II, of which their structure and functions in immune response are well defined, MHC class III are poorly defined structurally and functionally. It covers 700 kb and contains 61 genes, making it the most gene-dense region of the human genome. The functions of many genes are yet unknown. CYP21A2 travels in tandem with a pseudogene and the high degree of sequence similarity between them indicates that these two genes are evolving in tandem through intergenic exchange of DNA. CYP21A2 is located within the RCCX Cluster, which is the most complex gene cluster in the human genome. Which includes overlapping genes and genes within genes. In addition, containing a high density of retroelements such as human endogenous retrovirus and Alu elements. Like other cytochrome P450 enzymes, 21-hydroxylase participates in the cytochrome P450 catalytic cycle and engages in one-electron transfer with NADPH-P450 reductase. Its structure includes an essential iron heme group centered within the protein, also common to all P450 enzymes. Variations of the 21-hydroxylase enzyme can be found in all vertebrates. However, understanding of human 21-hydroxylase structure and function is of particular clinical value, as a failure of the enzyme to act appropriately results in congenital adrenal hyperplasia. The X-ray crystal structure for human 21-hydroxylase, with bound progesterone, was realized and published in 2015, providing opportunity for further study. The enzyme is notable for its substrate specificity and relatively high catalytic efficiency.
Structure
21-hydroxylase is a complex of three independent and identical enzyme subunits. Each subunit in the human enzyme consists of 13 ??α-helices and 9 ß-strands, formed into a triangular prism-like tertiary structure. The iron heme group that defines the active site resides in the center of each subunit. The human enzyme binds one substrate at a time. In contrast, the well-characterized bovine enzyme can bind two substrates. The human and bovine enzyme share 80% amino acid sequence identity, but are structurally different, particularly in loop regions, and also evident in secondary structure elements.
Function
This gene encodes a member of the cytochrome P450 superfamily of enzymes. The cytochrome P450 proteins are monooxygenases which catalyze many reactions involved in drug metabolism and synthesis of cholesterol, steroids and other lipids. This protein localizes to the endoplasmic reticulum and hydroxylates steroids at the 21 position. The 21-hydroxylase enzyme is one of three microsomal steroidogenic P450 enzymes, the others being 17-hydroxylase and aromatase. 21-hydroxylase is an essential enzyme in the biosynthetic pathways that produce cortisol and aldosterone.
Reaction
21-Hydroxylase catalyzes the addition of hydroxyl to the C21 position of two steroids: progesterone and 17α-hydroxyprogesterone.
Mechanism
21-Hydroxylase is a cytochrome P450 enzyme and follows the P450 catalytic cycle.
Kinetics
21-Hydroxylase is highly specific for hydroxylation of progesterone and 17-hydroxyprogesterone. No studies have reported sufficient binding of alternate substrates. In this way, it differs from the evolutionarily and functionally related P450 enzyme 17-hydroxylase, which has a large range of substrates. Earlier studies of the human enzyme expressed in yeast classified 17-hydroxyprogesterone as the best substrate for 21-hydroxylase. However, recent analysis of the purified human enzyme found a lower KM and greater catalytic efficiency for progesterone over 17-hydroxyprogesterone. The 2015 analysis found the catalytic efficiency of 21-hydroxylase for conversion of progesterone in humans to be approximately 1.3 x 10^7 M-1s-1 at 37 °C. This makes it the most catalytically efficient P450 enzyme of those reported, as of 2015, and more catalytically efficient than the closely related bovine 21-hydroxylase enzyme. C-H bond breaking to create a primary carbonradical is thought to be the rate-limiting step in the hydroxylation.
Congenital adrenal hyperplasia is an autosomal recessive disorder, and occurs in approximately 1 in 15000 births globally. There are multiple forms of CAH, broken down into classical and nonclassical forms based on the amount of function retained. The classical forms include salt-wasting, and simple-viralizing. Mutations that interfere with the active site—the heme group or residues involved in substrate binding—result in a complete loss of enzymatic activity, the salt-wasting type. Studies have demonstrated that mutations in 21-hydroxylase structure is related with clinical severity of congenital adrenal hyperplasia Cortisol and aldosterone deficits are associated with life-threatening salt-loss, as the steroids play roles in regulating sodiumhomeostasis. Retaining minimal enzyme activity, the simple-viralizing type is associated with mutations in conserved hydrophobic regions or near the transmembrane domain. Simple-viralizing CAH patients maintain adequate sodium homeostasis, but exhibit other phenotypical symptoms shared by SW, including accelerated growth in childhood and ambiguous genitalia in female neonates. Nonclassical forms retain 20-60% of hydroxylase function—this form is associated with normal cortisol expression, but an excess of androgens post-puberty.
Non-classic congenital adrenal hyperplasia
Non-classic congenital adrenal hyperplasia due to 21-hydroxylase deficiency is a milder and later-onset form of congenital adrenal hyperplasia. Some people affected by the condition have no associated signs and symptoms while others experience symptoms of androgen excess. Women with NCAH are generally born with normal female genitalia. Later in life, signs and symptoms of the condition can vary but may include hirsutism, frontal baldness, delayed menarche, menstrual irregularities, and infertility. Little has been published about males with NCAH. They may have early beard growth and relatively small testes. Typically, they have normal sperm counts. NCAH is caused by mutations in the CYP21A2 gene and is inherited in an autosomal recessive manner.
