Enzymology of butanol formation in Clostridium Beijerinckii

The present study encompasses an investigation of the expression of solvent forming enzymes and purification and characterization of butanol-forming enzymes. More sensitive and accurate procedures for the determination of acids and solvents in cultures have been developed, which led to the recognition of the onset of solvent production at the mid-exponential phase, about two h earlier than previously reported. Activities of solvent-forming enzymes started to increase about one h before the onset of measurable solvent production and the activities of solvent-forming enzymes did not increase simultaneously. CoA-acylating aldehyde dehydrogenase (ALDH) was purified to near homogeneity. The ALDH showed a native M.. of 100,000, and a subunit Mr of 55,000. ALDH could use either NAD(H) or NADP(H) as the coenzyme. ALDH was oxygenlabile. The O₂-inactivated enzyme could be reactivated by incubating the enzyme with CoA. Both NADH- and NADPH-dependent alcohol dehydrogenase activities were present in crude extracts. The ratio of NADPH-dependent activity to NADH-dependent activity (the PID ratio) varied in crude extracts. The PID ratio was affected by O~ ionic strength, pH, growth stage of cell, Fe in culture medium and temperature. Two ADHs have been identified in crude extracts. The NADPH-dependent ADH (P-ADH) could be separated from the NADH/NADPH-dependent ADH (D/P-ADH). The D/P-ADH has been extensively purified. The D/P-ADH showed a native Mr of 70,000 and subunits with Mr of 45,300 and 40,000. The D/P-ADH activity could be inactivated by a,a' -dipyridyl and restored by Fe2+.

Name thesis:degree_name

Ph. D.

Level thesis:degree_level

doctoral

Discipline thesis:degree_discipline

Anaerobic Microbiology

Department dc:contributor.department

Anaerobic Microbiology

Grantor dc:publisher

Virginia Tech

Year dc:date.issued

1991

Author dc:creator

• Yan, Run-Tao

Chair dc:contributor.committeechair

• Chen, Jiann-Shin

Committee members dc:contributor.committeemember

• Dean, Dennis R.
• Johnson, John L.
• Gregory, Eugene M.
• Krieg, Noel R.

Statement dc:rights

• In Copyright

Licence dc:rights.uri

• http://rightsstatements.org/vocab/InC/1.0/

Language dc:language.iso

en

Dc Identifier Other

etd-06192006-125711

OAI identifier oai:identifier

oai:vtechworks.lib.vt.edu:10919/38617