Sandbox Reserved 773

This Sandbox is Reserved from Sep 25, 2013, through Mar 31, 2014 for use in the course "BCH455/555 Proteins and Molecular Mechanisms" taught by Michael B. Goshe at the North Carolina State University. This reservation includes Sandbox Reserved 299, Sandbox Reserved 300 and Sandbox Reserved 760 through Sandbox Reserved 779.

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Histidine DecarboxylaseHistidine Decarboxylase

Figure 1. Asymmetrical unit of Histidine Decarboxylase formed by 3 homodimer-subunits)

Histidine Decarboxylase (HDC) is an enzyme that is responsible for converting histamine from amino acid L-histidine. This enzyme belong in the group II pyridoxal-5-phosphate (PLP)-dependent decarboxylase family . As the name suggested, this enzyme catalyzes the production of histamine by the removal of carboxylate group from the amino acid L-histidine whilst utilizes on pyridoxal phosphate as a cofactor . Since this enzyme breaks the carbon-carbon bond to produce carbon dioxide (CO2), it is in class IV lyase.

The mammalian Histamine decarboxylase is originated from HDC gene which in found in the human chromosome 15. This gene encodes a 74kDa precursor polypeptide . However, the enzyme becomes active after undergo post-translation proteolysis when its C-terminal is truncated into 54kDa .

Due to the fact that histamine is a mediator that triggers inflammation response associated with allergic reaction, HDC an important enzyme to study for the development of drugs or treatments for allergic disease .

General InformationGeneral Information

Histidine Decarboxylase

Figure 2. Sequence of Histidine Decarboxylase with its corresponding secondary stuctures

Symbol: HDC

Gene Name: HDC gene

Organism: Homo sapiens

Classification: Lyase

Length: 481 residues

Chains: A, B, C, D, E, F

Molecular Weight: 54314.8 kDa per chain

Isoelectric Point: 5.4 (mouse HDC)

Km: 0.1 mM (human) , 0.29mM (mouse stomach) , 0.26mM (mouse mastocytoma P-815 cells)

Vmax: 1880 nmol/min/mg

Sequence and StructureSequence and Structure

Histidine Decarboxylase is considered to be a homo-dimer when one observe its biological assembly . A homodimer is a quaternary structure formed by two identical monomers or protein chains. In human, three human HDC (hHDC) homodimers can be joined together to form a trimer in the asymmetric unit which can be seen in the 3D . Thus, one can use the nomenclature “trimer of dimer” to suggest the complex might dissociate into smaller subunits before dissociating into monomers. Specifically, and are found to account for the oligomerization process of the HDC complex.

Each monomer is divided into 3 structural . They are: the (~2-71), the (71-371), and the (372-477) (Figure 5) . A monomer is also composed of 49% and 13% (7 antiparallel and 4 parallel β-sheets). One distinctively long which span from Val-359 to Arg-393 connects the large and small domains together (Figure 2). Through hydrophobic effect, the N-terminal regions of the two monomers interact with each other extensively. At the same time, the large domains interact extensively due to electrostatic interactions. Thus, the N-terminal regions and large domains form the dimer interfaces of HDC .

spinqualitylabelsall models Asymmetric unit of Histidine Decarboxylase complex with 6 PLP-HME substrate-analogs (atoms shown in orange, grey, and blue) bound to each of the active sites. Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

Cofactor and Substrate Binding PocketCofactor and Substrate Binding Pocket

Figure 3. Interactions between enzyme HDC and cofactor-substrate PLP-HME at the binding site .

The for the cofactor PLP and the substrate histidine is located at the large domain. Since Histidine methyl ester (HME) is a substrate analog, PLP-HME can be utilized to demonstrate the binding interaction for the substrate-enzyme transition state at the active site . A one-dimensional representation of PLP-HME residing in the binding pocket can be seen in Figure 3. The of the active site are produced by several hydrophobic amino acids including Trp-72, Tyr-80, Leu-102, Phe-104, Ala-275, Tyr-334, Ile-436 (Figure 3). This hydrophobic pocket allow for the substrate to be protected from the solvent during the catalytic reaction.

The residue Val-150, Ser-151, Asp-302, Val-305, Ser-354 and a water molecule are all involved in the numerous hydrogen bonding to the phosphate group of PLP (Figure 3). More importantly, Ser-354 is found to be a critical residue for the substrate-binding pocket of HDC . Other residues that are within the hydrogen bonding distance to the substrate includes Tyr-81, His-194, Thr-248, and Asp-273 (Figure 3). The imidazole ring N1 and carbonyl O of HME are held in place by the side chains of Tyr-81 and His-194 respectively whereas the O and N in the pyridine ring of PLP are held in place by the side chains of Thr-248 and Asp-273 (Figure 3).

A loop region appears between the residues 330 to 340 of the large domain . This loop protrudes from the large domain into the active site of another subunit of the dimer and makes up the entrance of the active site. Especially Tyr-334, which directly interacts with the backbone of Ser-195 through a fairly weak hydrogen bond . Due to the weak hydrogen bonding, the loop’s position is not rigidly fixed. Thus, this causes this loop to be fairly flexible enabling it to act as a gate to “open” or “close” the active site .

Enzymatic MechanismEnzymatic Mechanism

During the catalytic reaction by HDC, the carboxyl group of L-histamine is removed with the help of cofactor pyridoxal-5'-phosphate (PLP) to generate the product histamine and byproduct carbon dioxide (CO2). The overall catalytic reaction is proposed to be a 1-step mechanism shown below :

Figure 4. Structure of HDC dimer with PLP and HME. Subunit A is shown in blue, Subunit B is shown in green, and the PLP-HME is shown in orange .

The substrate L-histidine is proposed to enter from the gate constructed by 5 hydrophobic residues (Tyr-80, Phe-104, Tyr-334, Leu-335, and Leu-353) along with PLP . This gate is compose of a very flexible loop structure which “closes” upon the entering of the substrate. This phenomenon could be explained by the hydrophobic effect, which compacts the flexible hydrophobic entrance towards the hydrophobic binding site, thereby trapping the substrate inside the catalytic region. As shown in Figure 3, the ester group of HME is located immediate to the entrance. Since the hydrophobic gate separates the solvent from the methyl ester of HME, it appears to facilitate the conversion of carboxylate group to the less hydrophilic CO2 product during catalysis .

The substrate specificity of HDC mechanism involves the residue Ser-354. Due to the Van der Waal effect of its hydroxyl side chain, Ser-354 avoids other 6-membered ring amino acid such as tyrosine from entering the substrate binding pocket. With this specificity, Ser-354 only allow for the 5-membered ring histidine to be involved in the binding site during catalysis .

The numerous hydrogen bond interaction between the enzyme and PLP restrict the translation of the substrate or cofactor. In addition, the negative charges of phosphate group of PLP is stabilized by dipole moment from the neighboring N-terminus of the helix α5 seen in Figure 4 . These hydrogen bonding and dipole moments create a stable environment for PLP to stay in place during the transitional state through proximity effect.

Additionally, the side chains of Thr-248 and Asp-273 are thought to be responsible for the protonation of the Oxide group and Nitrogen atom in the pyridine ring of PLP during the catalytic mechanism .

Pathways and ImplicationsPathways and Implications

Histidine Decarboxylase is a rather unstable enzyme with a short half-life of 50-100 min in varous conditions . This enzyme is only synthesized when histamine is needed and quickly degraded when enough histamine is made by Histamine N-methyltransferase (HMT) and Diamine oxidase (DAO) .

Furthermore, not all cells are found to make histamine. So far, only mask cells , basophils , neutrophil , enterochromafflin-like cells in gastric mucosa , and histaminergic neuron generate and store histamine. The histamine usually stored in the granula of these cells . Research have found that epithelial cell and lymphocytes can express HDC and synthesize histamine but cannot store them .

During a stimulation, the histamine are released in large amount which triggers many physiological process including:

- Allegoric reaction and Inflammation response

- Gastric acid secretion

- Capillary dilation and smooth muscle contraction

- Immune response

- neurotansmission in CNS

One of the most important role histamine play is the role of neurotransmitter in the central nervous system (CNS). When HDC is inhibited to function, it can change histaminergic neuron mediated physiological response listed below, as well as contributes to the probability for multiple diseases:

- Arousal

- Appetite

- Locomotion

- Tourette syndrome

- Schizophrenia

However outside of the central nervous system, anti-histamine drugs or HDC inhibitors are useful for treatment against allergic reaction, gastric ulcer, and inflammation