Sand box 326

Overview

3CBW exists as a homodimer quaternary structure.F Analyzing primary and quaternary structures of 3CBW with SPRITE and Chimera revealed two chains identical in both shape and sequence. Each chain is about 300 residues long, and the entire complex has a molecular weight of approximately 80.65 kDa.F

3CBW proteins originate from bacterial species.G,H InterPro search results show how nearly every enzyme with similar sequencing to 4Q7Q is found in various bacteria, with a notable exception to eukaryotes.D Additionally, the PDB entry for 4Q7Q notes how it potentially can be found in Chitinophaga pinensis, a gram-negative bacterial species which can degrade chitin.G,H

Family and Superfamily

Research shows that 4Q7Q is a member of the SGNH Hydrolase protein super family. BLAST and InterPro both suggested 4Q7Q’s inclusion in this family, and the known conserved residues seen from SPRITE analysis—Serine, Glycine, Asparagine, and Histidine—line up with those observed throughout this family.D,E Notably, this superfamily is also referred to as the GDSL Hydrolase superfamily.D,E

4Q7Q’s inclusion in this family also supports its SPRITE-derived hypothetical functionality. Rhamnogalacturonan Acetylesterase—the enzyme with one of the best SPRITE-based alignment relative to 4Q7Q—is a member of this family.F Proteins in this family are also known for containing a “unique hydrogen bond network that [stabilizes]” the active site.F

Regarding what protein family 4Q7Q belongs to, DALI results suggest it is a part of a sub-family of the greater GDSL/SGNH superfamily. A PDB90% DALI search labels 4Q7Q as a part of the “Lipolytic Protein G-D-S-L Family,” which refers to enzymes that hydrolyze lipid substrates.I

Sequence Analysis

The primary sequence of 4Q7Q shows several conserved sequences between it and esterase-like proteins. A sequence of GDSI—similar to the GDSL sequence seen from its family and superfamily—can be seen between 4Q7Q and enzymes like Isoamyl Acetate-Hydrolyzing Esterase. Other noteworthy conserved sequences between esterases and 4Q7Q include GxND and DGxH.

These enzymes also share similar secondary structures. Segments of alpha-helixes and beta-sheet strands appear and remain nearly entirely conserved throughout esterase analysis. A few conserved coils appear, but these sections do not appear as often as the other two secondary structures.

Similar conserved sequences could be found between 4Q7Q and lipases. The GDSI, GxND, and DGxH sequences can be seen from lipases like 7BXD.? The same secondary structure segments can also be located in the lipases analyzed.

Structural Analysis (DONE)

Using Right-hand SPRITE analysis, it showed that 3BCW had similar residues to small amino acid chains. Specific residues are Ala. 76, Ile. 79, Gly. 61, Val. 293, the first two being on the A chain side and the second being on the B chain side of the 3CBW. Comparing 3CBW to 1BHG, which is human beta-glucuronidase, the RMSD value was a difference of 0.46 angstroms.

With the use of DALI, it was determined that 3CBW is most similar to Beta-1,4-Mannanase and Mannan Endo-1,4-Beta-Mannosidase. Both of these enzymes hydrolyzes different linkages.

Substrates

Using SwissDock, it was determined that larger, cyclic molecules are good substrates that would have the best binding affinity to 3CBW, each had about a -7 kcal/mol binding affinity. These large molecules are 4-Nitrophenyl N-acetyl-β-D-glucosaminide, 4-Nitrophenyl α-D-glucopyranoside, and p-nitrophenyl phosphate.

Despite this, these ligands show noticeable hydrophobic interactions with the active site. This implies 4Q7Q uses hydrophobic regions to help guide substrates into the right orientation for enzymatic processes. This also further supports the possibility that 4Q7Q primarily operates with hydrophobic lipid-based substrates. This also explains why Methyl Acetate exhibited a relatively weaker affinity for 4Q7Q, as its smaller structure prevented hydrophobic interactions.

Theoretical Functionality and Proposed Bodily Purpose

Highlight the data that helped you come to your conclusion here including any relevant figures. Make sure include potential substrates and binding sites.

Potential Substrates and Binding Sites: 4-Nitrophenyl N-acetyl-β-D-glucosaminide, 4-Nitrophenyl α-D-glucopyranoside, and P-Nitrophenyl Phosphate