User:Betaclamp/Sandbox

DNA polymerase III subunit beta
Crystallographic structure of the dimeric DNA polymerase beta subunit from E. coli.
Identifiers
Organism	Escherichia coli
Symbol	dnaN
Entrez	948218
PDB	1MMI
RefSeq (Prot)	NP_418156
UniProt	P0A988
Other data
EC number	2.7.7.7
Chromosome	MG1655: 3.88 - 3.88 Mb
Search for Structures Swiss-model Domains InterPro

The beta clamp is a type of DNA clamp found in prokaryotes, that ensnares DNA and then allows a point-of-attachment for DNA replicating enzymes. The beta clamp is a dimeric subunit of prokaryotic DNA polymerase III which acts as a sliding clamp to keep the polymerase bound to the DNA. DNA polymerase III is the primary enzyme complex involved in both prokaryotic and eukaryotic DNA replication.

The gamma complex of DNA polymerase III (composed of γδδ'χψ subunits) hydrolyzes ATP to chaperone two beta subunits to bind to DNA. Once bound to DNA, the beta subunits can freely slide along double stranded DNA. The complex between the dimeric beta protein and DNA is known as the pre-initiation complex. The beta subunits in turn bind the αε polymerase complex (where the α subunit possesses DNA polymerase activity and the ε subunit is a 3‘-5’ exonuclease).

Mechanism

A spliceosome is sized at over a million kD. Also based in the cell nucleus are multitudes of massive biomolecular constructs - helicases, isomerases, Dna replicases and Rna polymerases - all massive constructs. In perspective, a DNA double strand is only 20 atoms across [in diameter].

One DNA molecule (times 23 pairs) is at the constant beck-and-call of the dozens of types (not copy numbers) of these highly variant, space and raw materials requiring giants and requires an intermediary that binds the DNA and then proffers its other active binding site that will then be approached by and utilized as a point of attachment to the 'factoryase'. That intermediary is the beta clamp which consists of two parts in the prokaryotes, while its counterpart in eukaryotes is termed PCNA and is a trimer.

Being composed of subunits, the beta clamp is naturally itself a subunit - a component of a construct that, when fully assembled, is called a holoenzyme. For illustration, the replicase DNA polymerase III is a factoryase that is concerned with the replication ( copying via duplication ) of double stranded (ds) DNA. Its capabilities include being able to replicate both strands, in opposite directions, simultaneously (see semi-discontinuous replication).

It comprises nine types ( molecular species ) of subunits : the alpha - DNA synthesis (130 kD), epsilon - proof reading (25 kD) and theta - assembly? (10 kD) make up a catalytic core. Tau (71 kD) causes two of these cores to assemble and bind, i.e. dimerise, forming Pol III* . Addition of gamma (55 kD), { which itself has several subunits } , forms Pol III' (750 kD). Gamma and delta (32 kD) together bind to the DNA template at a control sequence. A pair of beta subunits form the Beta Dimer that along with two other subunits complete the holoenzyme ( 900 kD ).

DNA Pol III subunit inventory, as fully assembled :

First core = 2 beta, 2 delta, 1 gamma, 1 alpha, 1 epsilon, 1 theta, 1 psi and 1 chi

Second core = 2 beta, 2 tau (to hold the two alphas together), 1 alpha, 1 epsilon, and 1 theta .

Holoenzyme total = 2 alpha, 4 beta, 1 gamma, 2 delta, 2 epsilon, 2 theta, 2 tau, 1 psi and 1 chi totals 17 subunits .

The structure of DNA Pol III is assembled in three stages.

1. Assembly begins with the coalescence of the five-subunit 'clamp loader' (comprised of two delta subunits, one gamma subunit, one psi subunit and one chi subunit). This 'gamma complex' binds with a beta dimer forming a pre-initiation complex. This complex hydrolyzes ATP to complete the encirclement of the DNA duplex molecule.
2. As always, intermolecular binding results in conformational changes. For example : the Kinase class of factoryases uses the docking and undocking of just one phosphate moiety to alter the shape of its bulk to switch between active and inactive forms. When the beta dimer clamps the DNA, its affinity for the gamma complex is replaced by an attraction for the core-polymerase, which brings the core to DNA.
3. A pair of Tau(s) dimerize and bind to the first core; the second Tau captures the second core along with its beta clamp.

When DNA polymerase has completed replicating a stretch of DNA, the polymerase dislocates from the template strand. The sliding clamp, however, remains in place for other proteins to use.

Structure

DNA polymerase III, beta chain Identifiers Symbol DNA_polIII_beta Pfam PF00712 InterPro IPR001001 SMART SM00480 SCOP2 2pol / SCOPe / SUPFAM Available protein structures: Pfam   structures / ECOD   PDB RCSB PDB; PDBe; PDBj PDBsum structure summary PDB 1jqj​

DNA polymerase III, beta chain, N-terminal Identifiers Symbol DNA_pol3_beta Pfam PF00712 InterPro IPR022634 Available protein structures: Pfam   structures / ECOD   PDB RCSB PDB; PDBe; PDBj PDBsum structure summary

DNA polymerase III, beta chain, central Identifiers Symbol DNA_pol3_beta_2 Pfam PF02767 InterPro IPR022637 Available protein structures: Pfam   structures / ECOD   PDB RCSB PDB; PDBe; PDBj PDBsum structure summary

DNA polymerase III, beta chain, C-terminal Identifiers Symbol DNA_pol3_beta_3 Pfam PF02768 InterPro IPR022635 Available protein structures: Pfam   structures / ECOD   PDB RCSB PDB; PDBe; PDBj PDBsum structure summary

Crystallographic studies of beta clamp shows that it forms a ring shaped dimer, the beta ring. This beta ring clamp empowers the holoenzyme with very fast procession speeds. The beta clamp completely surrounds the DNA, yet can easily slide along a DNA duplex.

Honoring B. Lewin's contribution, here is an unparaphrasable passage:

The dimer surrounds the duplex, providing the "sliding clamp" that allows the holoenzyme to slide along DNA. The structure explains the high processivity - there is no way for the enzyme to fall off !

Processivity

In the absence of a sliding clamp, DNA polymerase has a processivity of only 20 to 100 bp. That is to say, the DNA polymerase would fall off of the template strand every 20 to 100 bp, dramatically slowing down DNA replication. With the clamp, the polymerase still falls off frequently, but remains anchored to the template strand and can resume polymerization much faster than were it allowed to diffuse away.

As an application and exercise to focus on quantities : the Escherichia coli bacteria's genetic complement consists of one double-stranded Dna chromosome that exists, at least during replication, in a closed-loop [ or circular ] conformation, and is comprised of 4.2 million base pairs.

Under optimal growth conditions, E. coli will replicate its chromosome in approximately 42 minutes.

The chromosome being a circle, copying must begin somewhere : this point of origin of replication is not selected randomly, but occurs at a fixed recognition sequence in the Dna named oriC . The main point here is that when replication begins at this origin, it proceeds using two polymerases, one each in the opposite direction, thereby cutting the replication time in half. Therefore, a correction factor of 0.5 must be added to the following calculation.

Dividing quantity by time = 4,200,000 bp / 42 minutes * 0.5 = 50,000 base pairs replicated per minute { Genes VI Table 14.1 pg 437 } or 833 base pairs per second !

Another recent discovery also highlights the speeds involved. For Dna to be copied, the two strands must first be unwound and separated. The helicase class of enzymes that perform this function have parts that spin at 10,000 revolutions per minute, or 167 revolutions per second.

See also

• dnaN
• DNA polymerase I
• DNA polymerase II

Further reading

• Lewin, Benjamin (1997). Genes VI. Oxford [Oxfordshire]: Oxford University Press. pp. 484–7. ISBN 0-19-857779-6.

External links

• beta+subunit,+DNA+polymerase+III at the U.S. National Library of Medicine Medical Subject Headings (MeSH)

Category:Molecular genetics