A pathogenic linked mutation in the catalytic core of human cystathionine beta-synthase disrupts allosteric regulation and allows kinetic characterization of a full-length dimer.

A pathogenic linked mutation in the catalytic core of human cystathionine beta-synthase disrupts allosteric regulation and allows kinetic characterization of a full-length dimer. Research Abstract Details 

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A pathogenic linked mutation in the catalytic core of human cystathionine beta-synthase disrupts allosteric regulation and allows kinetic characterization of a full-length dimer. Abstract Text:

Suvajit Sen,Ruma Banerjee,

Cystathionine beta-synthase catalyzes the condensation of serine and homocysteine to yield cystathionine and is the single most common locus of mutations associated with homocystinuria. In this study, we have examined the kinetic consequences of a pair of linked patient mutations, P78R/K102N, that are housed in the catalytic core of the protein and compared it to the effects of the corresponding single mutations. The P78R mutation affords purification of a mixture of higher order oligomers, P78R-I, which resembles the mixed quaternary state associated with wild-type enzyme. However, unlike wild-type enzyme, P78R-I converts over time to P78R-II, which exists predominantly as a full-length dimer. The specific activities of the K102N, P78R-I, and P78R-II mutants in the absence of AdoMet are approximately 3-, 9-, and 3-fold lower than of wild-type enzyme and are stimulated 2.9-, 2.5-, and 1.4-fold respectively by AdoMet. However, when linked, the specific activity of the resulting double mutant is comparable to that of wild-type enzyme but it is unresponsive to AdoMet, revealing that interactions between the two sites modulate the phenotype of the enzyme. Steady-state kinetic analysis for the double mutant reveals a sigmoidal dependence on homocysteine that is not observed with wild-type enzyme, which is ascribed to the mutation at the K102 locus and indicates changes in subunit interactions. Hydrogen-deuterium mass spectrometric analysis reveals that, even in the absence of AdoMet, the double mutant is locked in an activated conformation that is observed for wild-type enzyme in the presence of AdoMet, providing a structural rationale for loss of this allosteric regulation. To our knowledge, this is the first example of mutations in the catalytic core of cystathionine beta-synthase that result in failure of AdoMet-dependent regulation. Furthermore, analysis of individual single mutations has permitted, for the first time, partial kinetic characterization of a full-length dimeric form of human cystathionine beta-synthase.

A pathogenic linked mutation in the catalytic core of human cystathionine beta-synthase disrupts allosteric regulation and allows kinetic characterization of a full-length dimer. Publishing Authors By Initials

S Sen,R Banerjee,

For similar amino acids, sulfur: methionine: s-adenosylmethionine research abstracts see: amino acids, sulfur: methionine: s-adenosylmethionine research

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A pathogenic linked mutation in the catalytic core of human cystathionine beta-synthase disrupts allosteric regulation and allows kinetic characterization of a full-length dimer. Journal Published:

PUBLICATION TYPE: Research Support, N.I.H., Extr

Journal: Biochemistry

VOLUME: 46

Page Numbers: 4110-6

Journal Abbreviation: Biochemistry

ISSN: 0006-2960

DAY: 13

MONTH: 03

YEAR: 2007

A pathogenic linked mutation in the catalytic core of human cystathionine beta-synthase disrupts allosteric regulation and allows kinetic characterization of a full-length dimer. Information

Number of References:

LANGUAGE: eng

NlmUniqueID: 370623

A pathogenic linked mutation in the catalytic core of human cystathionine beta-synthase disrupts allosteric regulation and allows kinetic characterization of a full-length dimer. Keywords Mesh Terms:

KEYWORDS: S-Adenosylmethionine

MESH TERMS: physiology

Chemical & Substance for Abstract: A pathogenic linked mutation in the catalytic core of human cystathionine beta-synthase disrupts allosteric regulation and allows kinetic characterization of a full-length dimer. Information

Substance Name: Cystathionine beta-Synthase

Registry Number: EC 4.2.1.22

Grant and Affiliation Information for A pathogenic linked mutation in the catalytic core of human cystathionine beta-synthase disrupts allosteric regulation and allows kinetic characterization of a full-length dimer.

AFFILIATION: Redox Biology Center and the Biochemistry Department, University of Nebraska, Lincoln, Nebraska 68588-0664, USA.

Country: United States

AGENCY: United States NCRR

GRANT: P20RR17675

ACRONYM: RR

MEDLINETA: Biochemistry

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